Antigens and related assay to detect and measure auto-antibodies in a subject developing type 1 diabetes (T1D)

Abstract

The invention relates to a diagnostic kit and methods for detecting and quantifying autoantibodies associated with the onset of type 1 diabetes (T1D). The kit comprises recombinant antigen molecules fused to bioluminescent reporter proteins for the detection of autoantibodies directed against self-antigens expressed by pancreatic islet beta cells. The presence, absence, or amount of one or more autoantibody molecules against said self-antigens is determined with a luciferase immune precipitation system (LIPS) assay or a modified version of a LIPS assay, i.e., a solid phase capture LIPS (scLIPS) assay.

Description

[0001] Antigens and related assay to detect and measure auto-antibodies in a subject developing type 1 diabetes (T1D)

[0002] DESCRIPTION FIELD OF THE INVENTION

[0003] The present invention relates to a test kit for performing an assay to detect and measure at least one autoantibody molecule in a subject developing type 1 diabetes (T1D). The invention further relates to methods for determining the presence and / or level of at least one autoantibody molecule in a sample from a human subject developing T1D using said recombinant antigen molecules.

[0004] BACKGROUND

[0005] Type 1 diabetes (T1D) is an autoimmune disease characterized by the progressive destruction of pancreatic p-cells, the only cells responsible for insulin production. Clinical onset is often acute and associated with significant complications, particularly in children1. Despite a century of availability, insulin therapy remains a replacement treatment rather than a cure; therefore, preventing or halting the autoimmune response against pancreatic islets remains a central goal. Four major autoantigens are targeted by the immune system in individuals at risk for or affected by T1D: insulin / proinsulin, GAD65, IA-2, and ZnT8. The corresponding autoantibodies (IAA, GADA, IA-2A, ZnT8A) are essential biomarkers for diagnosis2, prediction of disease progression3, and monitoring of experimental preventive therapies4’56. Large screening and surveillance programs (e.g., Frida, ASK, TEDDY) require the execution of hundreds of thousands of tests in pediatric and general populations. Early identification of individuals positive for multiple autoantibodies is crucial, as multiple positivity strongly correlates with progression to overt T1 D7’8’9’10

[0006] Islet autoantibodies typically exhibit low titers and specificity for conformational epitopes, necessitating highly sensitive assays that preserve antigen structure. For this reason, Radio Binding Assays (RBAs) have become the gold standard. However, the use of radioisotopes imposes significant limitations: stringent regulatory requirements, high costs, logistical burdens, and uncertain long-term sustainability for large-scale population screening.

[0007] Several non-radioactive formats have been proposed, including Bridge-ELISA14, ECL181920, and ADAP21, showing overall good performance but with important drawbacks: poor detection of IAA with conventional ELISAs15, high serum volume requirements, need for dedicated platforms, and limited compatibility with high-throughput diagnostic workflows11■121 13. These weaknesses are especially problematic in programs relying on low-volume capillary samples.

[0008] The Luciferase Immunoprecipitation System (LIPS) is conceptually similar to RBA but avoids radioactivity. Classical LIPS assays based on Firefly or Renilla luciferases have demonstrated utility for GADA and IA-2A detection. More recently, the small and highly processive NanoLuc™24reporter luciferase has been used to improve LIPS sensitivity for autoantibodies against ZnT825, GAD6526, and insulin27. Despite these advances, assay performance — particularly specificity anddisease-discrimination capability — still varies among platforms, and the need for high specificity is especially critical in low-prevalence population screening.

[0009] There remains an unmet need for robust, highly sensitive and specific, low-cost, high-throughput assays with minimal sample requirements and full compatibility with automated diagnostic platforms. Such assays are essential for expanding general-population screening initiatives and enabling new longitudinal studies of T1D autoimmunity.

[0010] SUMMARY OF THE INVENTION

[0011] The present invention addresses the above-mentioned needs and provides a test kit for performing an assay, the assay comprising a set of at least three recombinant antigen molecules in a single composition. In a preferred aspect, the antigen molecules are selected from the group comprising recombinant antigen molecules selected from the group comprising recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin, and recombinant human proinsulin. In another preferred aspect, the antigen molecules are selected from the group consisting of recombinant antigen molecules selected from the group comprising recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin, and recombinant human proinsulin. In one further aspect, the recombinant antigen molecules are fused to a bioluminescent reporter protein. According to the invention, the test kit also comprises a substrate for the bioluminescent reporter protein.

[0012] The recombinant antigen molecules according to the invention can be used for the diagnosis of type 1 diabetes in human patients. It is expected that said use of the recombinant antigen molecules will improve the rapidity and the accuracy of detection of islet autoantibodies due to the assay multiplexing, their specificity, and sensitivity.

[0013] Thus, the recombinant antigen molecules according to the invention are particularly suitable for binding to an autoantibody molecule, specifically an islet autoantibody molecule, more specifically an autoantibody with specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, or human Zinc transporter 8, further optionally wherein the antibody molecule is at least one islet autoantibody selected from the group comprising islet autoantibodies IAA (Insulin Auto Antibodies), GADA (Glutamic Acid Decarboxylase Antibodies), IA-2A (Islet Antigen 2 Antibodies), and ZnT8A (Zinc Transporter 8 Antibodies). Preferably, the binding of the autoantibody molecule to at least one recombinant antigen molecule according to the invention is binding in a liquid phase detection assay. This is particularly advantageouscompared to other detection assays, e.g., such as ELISA, as the structure of the recombinant antigen, i.e., the tertiary structure of the recombinant antigen, is preserved.

[0014] The recombinant antigen molecules according to the invention can be used in a luciferase immune precipitation system (LIPS) assay or a modified version of the LIPS assay, herein referred to as a solid phase capture LIPS (scLIPS) assay.

[0015] The present invention further provides a set of nucleic acids encoding the recombinant antigen molecules of the test kit. Preferably, each recombinant antigen molecule is encoded by a separate nucleic acid.

[0016] The present invention also relates a vector encoding for:

[0017] a) a recombinant human insulin antigen molecule encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1, 2, 10, 11, and 22, preferably, encoded by a sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 1 or 2;

[0018] b) a recombinant human proinsulin antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 8 and 9;

[0019] c) a recombinant human Glutamate decarboxylase 65 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 3, 7, and 16 to 18, preferably encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to SEQ ID NO: 3;

[0020] d) a recombinant human islet antigen 2 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 4, 14, 15, and 19, preferably encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to SEQ ID NO: 4; or

[0021] e) a recombinant human Zinc transporter 8 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 5, 6, 12, 13, and 20, preferably encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to SEQ ID NOs: 5 or 6.

[0022] In addition, the present invention provides a cell comprising the set of nucleic acids, or a nucleic acid, or a vector, and expressing the recombinant antigen molecules. In one embodiment, the recombinant antigen coding sequence is inserted into a lentiviral vector. Preferably, the cell is a stably expressing transfectant cell line, such as an Expi-293F cell line.Additionally, the present invention also provides a method for producing the recombinant antigen molecules according to the present invention, the method comprising expressing said nucleic acids or said vector in a cell according to the present invention, and harvesting said recombinant antigen molecules.

[0023] In a further aspect, the present invention provides an in vitro method for determining the presence and / or level of an antibody molecule in a sample. In one aspect, the sample is a blood sample, a serum sample, or a plasma sample obtained from a human subject.

[0024] The in vitro method comprises the steps of:

[0025] a) contacting the sample with the set of at least three recombinant antigen molecules, wherein a complex between the recombinant antigen molecule and the autoantibody molecule (referred to as “antigen molecule-autoantibody complex”) is formed;

[0026] b) removing the unbound recombinant antigen molecules;

[0027] c) contacting the recombinant antigen molecule-autoantibody complex with a protein; d) contacting the bioluminescence reporter protein covalently linked to the recombinant antigen molecule with the substrate; and

[0028] e) determining the presence, absence, and / or amount of one or more autoantibody molecules in the sample that binds to one or more recombinant antigen molecules.

[0029] The presence, absence, or amount of one or more autoantibody molecules in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter molecule. The at least one autoantibody molecule is against a self-antigen. Preferably, the at least one autoantibody molecule is an islet autoantibody molecule. More preferably, the islet autoantibody molecule has specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, or human Zinc transporter 8. Most preferably, the islet autoantibody molecule is selected from the group comprising or consisting of islet autoantibodies IAA (Insulin Auto Antibodies), GADA (Glutamic Acid Decarboxylase Antibodies), IA-2A (Islet Antigen 2 Antibodies), and ZnT8A (Zinc Transporter 8 Antibodies).

[0030] In a first aspect, the invention relates to an in vitro method for determining the presence and / or level of at least one autoantibody in a sample, wherein the sample is a blood sample, a serum sample, or a plasma sample obtained from a human subject, the method comprising:

[0031] a) contacting the sample with at least three recombinant antigens covalently linked to a bioluminescence reporter and forming at least one complex between the at least three recombinant antigens and the at least one autoantibody,

[0032] b) contacting the complex formed in step a) with at least one protein selected from the group comprising or consisting of protein A, protein G, protein AG, protein L, anti IgA, anti IgM, and anti IgG; andc) determining the presence, absence, and / or amount of the at least one autoantibody in the sample that binds to the at least three recombinant antigens,

[0033] wherein the at least three recombinant antigens are selected from the group consisting of: recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin and recombinant human proinsulin, and

[0034] wherein the presence, absence, or amount of at least one autoantibody in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter.

[0035] Preferably, the presence of at least one autoantibody that binds to at least one of the at least three recombinant antigens in the sample is indicative of type 1 diabetes in the human subject or of a risk of developing type 1 diabetes.

[0036] Preferably, the presence in the sample of at least two autoantibodies binding to at least two different recombinant antigens among the at least three recombinant antigens are indicative of type 1 diabetes in the human subject or of a risk of developing type 1 diabetes.

[0037] Preferably, the presence in the sample of at least three autoantibodies binding to the at least three recombinant antigens are indicative of type 1 diabetes in the human subject or of a risk of developing type 1 diabetes.

[0038] In some embodiments, the at least three recombinant antigens are Glutamate decarboxylase 65, recombinant human islet antigen 2 and recombinant human Zinc transporter 8. Preferably, the method is performed by a luciferase immune precipitation system (LIPS) assay, more preferably, the method is performed by a solid phase capture luciferase immune precipitation system (scLIPS) assay. In some embodiments, the at least three antigens do not comprise human insulin. In other embodiments, when at least one autoantibody is present in the sample, the sample is further contacted with recombinant human insulin.

[0039] In some embodiments, the sample is a dry sample that is reconstituted prior to the step of contacting the sample with the at least three antigens. Preferably, the sample is a dried blood spot (DBS).

[0040] A second aspect of the present invention relates to a kit for detecting autoantibodies associated with type 1 diabetes, comprising:

[0041] a) at least three recombinant antigens selected from the group comprising or consisting of: recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin, and recombinant human proinsulin, wherein each of the recombinant antigen molecules is fused to a bioluminescent reporter, and

[0042] b) a substrate for the bioluminescent reporter.In some embodiments, the kit comprises one or more compositions comprising the at least three antigens.

[0043] In some other embodiments, the kit comprises one composition comprising the at least three antigens.

[0044] Preferably, the composition comprises recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2 and recombinant human Zinc transporter 8.

[0045] Preferably, the bioluminescent reporter is a nanoluciferase reporter protein, more preferably, the nanoluciferase reporter protein is a Nanoluc luciferase (NLuc) reporter protein, even more preferably, the Nanoluc luciferase (NLuc) reporter protein is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99, or 100% identical to SEQ ID NO: 10.

[0046] In some embodiments, the assay is a luciferase immune precipitation system (LIPS) assay or a solid phase capture LIPS (scLIPS) assay.

[0047] Preferably, the kit comprises a protein selected from the group comprising or consisting of protein A, protein G, protein AG, protein L, anti IgA, anti IgM, and anti IgG.

[0048] Preferably:

[0049] a) the recombinant human insulin antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ I D NOs: 1, and 11 to 46;

[0050] b) the recombinant human proinsulin antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 65 to 87 and 96;

[0051] c) the recombinant human islet antigen 2 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 2 to 4;

[0052] d) the recombinant human Zinc transporter 8 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 5; and / or

[0053] e) the recombinant human Glutamate decarboxylase 65 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 6.

[0054] A third aspect of the invention relates to a vector comprising at least one nucleic acid selected from the group consisting of: SEQ ID NOs: 1, 11 to 46, SEQ ID NOs: 65 to 87, 96, SEQ ID NOs: 2 to 4, SEQ ID NO: 5 and SEQ ID NO: 6 or a sequence at least 85, 90, 95, 98, 99 or 100% identical to said sequences.

[0055] A fourth aspect of the present invention relates to a cell comprising the vector as defined above and expressing at least one of the at least three recombinant antigens. Preferably, the cell is astably expressing transfectant cell line, more preferably, the transfectant cell line is an Expi-293F cell line.

[0056] A fifth aspect of the present invention relates to method for producing the recombinant antigens of the test kit as defined above, the method comprising expressing the vector as defined above in a cell as defined above and harvesting the recombinant antigen molecules.

[0057] The invention is illustrated by means of non-limiting examples referring to the following figures:

[0058] BRIEF DESCRIPTION OF THE FIGURES

[0059] Figure 1: GADA LIPS and scLIPS performance in the IASP2023 interlaboratory comparison study. (A) Distribution of arbitrary units in case and control samples in GADA LIPS. GADA arbitrary units in blood donors (control, n = 90) and new onset T1 D patients (T 1 D, n= 50) samples. The violin plots correspond to their probability density estimate. (B) GADA LIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (C) GADA assay ROC curve in LIPS. Shown lines correspond to: GADA LIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (D) Distribution of arbitrary units in case and control samples in GADA scLIPS. GADA arbitrary units in blood donors (control, n = 90) and new onset T1D patients (T1D, n= 50) samples. The violin plots correspond to their probability density estimate. (E) GADA scLIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (F) GADA assay ROC curve in scLIPS. Shown lines correspond to: GADA scLIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (G) Boxplot of assay sensitivity and specificity according to positive scores assigned by laboratories. (H) Boxplot of ROC-pAUC95 across all submitted assays. (I) Boxplot and tilemap of positive scores assigned to each sample across all submitted assays. Figure 2: IA-2A LIPS and scLIPS performances in the IASP2023 interlaboratory comparison study. (A) Distribution of arbitrary units in case and control samples in IA-2A LIPS. IA-2A arbitrary units in blood donors (control, n = 90) and new onset T1 D patients (T 1 D, n= 50) samples. The violin plots correspond to their probability density estimate. (B) IA-2A LIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (C) IA-2A assay ROC curve in LIPS. Shown lines correspond to: IA-2A LIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (D) Distribution of arbitrary units in case and control samples in IA-2A scLIPS. IA-2A arbitrary units in blood donors (control, n = 90) and newonset T1D patients (T1D, n= 50) samples. The violin plots correspond to their probability density estimate. (E) IA-2A scLIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1 D patients (T 1 D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (F) IA-2A assay ROC curve in scLIPS. Shown lines correspond to: IA-2A scLIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (G) Boxplot of assay sensitivity and specificity according to positive scores assigned by laboratories. (H) Boxplot of ROC-pAUC95 across all submitted assays. (I) Boxplot and tilemap of positive scores assigned to each sample across all submitted assays.

[0060] Figure 3: ZnT8A LIPS and scLIPS performances in the IASP2023 interlaboratory comparison study. (A) Distribution of arbitrary units in case and control samples. ZnT8A arbitrary units in blood donors (control, n = 90) and new onset T1 D patients (T 1 D, n= 50) samples. The violin plots correspond to their probability density estimate. (B) ZnT8A LIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (C) ZnT8A assay ROC curve.

[0061] Shown lines correspond to: ZnT8A LIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (D) Distribution of arbitrary units in case and control samples. ZnT8A arbitrary units in blood donors (control, n = 90) and new onset T1D patients (T1D, n= 50) samples. The violin plots correspond to their probability density estimate.

[0062] (E) ZnT8A scLIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (F) ZnT8A assay ROC curve. Shown lines correspond to: ZnT8A scLIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (G) Boxplot of assay sensitivity and specificity across the submitted assay formats. (H) Boxplot of ROC-pAUC95 across all submitted assays. (I) Boxplot and tilemap of positive scores assigned to each sample across all submitted assays.

[0063] Figure 4: IAA LIPS and scLIPS performances in the IASP2023 interlaboratory comparison study. (A) Distribution of arbitrary units in case and control samples. IAA arbitrary units in blood donors (control, n = 90) and new onset T1 D patients (T 1 D, n= 50) samples. The violin plots correspond to their probability density estimate. (B) Insulin LIPS sensitivity and specificity.

[0064] Percent specificity and sensitivity calculated in blood donors (control) and new onset T1 D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (C) Insulin assay ROC curve. Shown lines correspond to: Insulin LIPS (grey lines); median ROC curve across all submitted assays (dottedline); identity line (dashed line). (D) Distribution of arbitrary units in case and control samples.

[0065] IAA arbitrary units in blood donors (control, n = 90) and new onset T1D patients (T1D, n= 50) samples. The violin plots correspond to their probability density estimate. (E) Insulin scLIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (F) Insulin assay ROC curve. Shown lines correspond to: Insulin scLIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (G) Boxplot of assay sensitivity and specificity across the submitted assay formats. (H) Boxplot of ROC-pAUC95 across all submitted assays. (I) Boxplot and tilemap of positive scores assigned to each sample across all submitted assays.

[0066] Figure 5: Multiplexed LIPS performance pilot experiment optimization of antigen input in the assay. (A) The plots show raw signal (light units) detected after pilot multiplexed LIPS assays in blood donors (n = 8) and new onset T1D patients (n= 14) samples. The assays differed by the amount of each GAD, IA-2 and ZnT8 antigens that were multiplexed: 4 million light units equivalents of each antigen i.e., like in single LIPS assays and 2 million light units equivalents of each antigen, respectively. The violin plots show the corresponding probability density estimates.

[0067] (B) Distribution of signal to noise ratio in case and control samples. The plots show the signal to noise relative to a negative control reference serum detected after pilot multiplexed LIPS assays in blood donors (n = 8) and new onset T1D patients (n= 14) samples. The assays differed by the amount of each GAD, IA-2 and ZnT8 antigens that were multiplexed: 4 million light units equivalents of each antigen i.e., like in single LIPS assays and 2 million light units equivalents of each antigen, respectively. The violin plots show the corresponding probability density estimates.

[0068] Figure 6: Optimization of antigen input in multiplexed LIPS leads to improved signal to noise ratio in control samples. (A) The plots show raw signal (light units, left panel) and signal to noise relative a negative control reference serum (right panel) in a pilot 3-screen LIPS in which different amounts of each GAD, IA-2 and ZnT8 antigens were mixed. The violin plots show the corresponding probability density estimates.

[0069] Figure 7: Multiplexed LIPS performance in discriminating case from control samples from the IASP2020 interlaboratory comparison study. (A) Distribution of arbitrary units in case and control samples. Multiplexed arbitrary units in blood donors (control, n = 90) and new onset T1D patients (T1D, n= 50) samples. The violin plots correspond to their probability density estimate.

[0070] (B) Multiplexed assay ROC curve. Shown lines correspond to: Multiplexed LIPS (black line); identity line (dashed line).

[0071] Figure 8: Multiplexed LIPS and scLIPS performances in discriminating case from control samples tested in blind in the IASP2023 interlaboratory comparison study. (A) Distributionof arbitrary units in case and control samples. Multiplexed arbitrary units in blood donors (control, n = 90) and new onset T1D patients (T1D, n= 50) samples. The violin plots correspond to their probability density estimate. (B) Multiplexed LIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (C) Multiplexed assay ROC curve. Shown lines correspond to: Multiplexed LIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (D) Distribution of arbitrary units in case and control samples. Multiplexed arbitrary units in blood donors (control, n = 90) and new onset T1D patients (T1D, n= 50) samples. The violin plots correspond to their probability density estimate. (E) Multiplexed scLIPS sensitivity and specificity. Percent specificity and sensitivity calculated in blood donors (control) and new onset T1D patients (T1D) samples. The dashed and dotted lines correspond to the median sensitivity and median specificity across all participating assay, respectively. (F) Multiplexed assay ROC curve. Shown lines correspond to: Multiplexed scLIPS (grey lines); median ROC curve across all submitted assays (dotted line); identity line (dashed line). (G) Boxplot of assay sensitivity and specificity across the submitted assay formats. (H) Boxplot of ROC-pAUC95 across all submitted assays. (I) Boxplot and tilemap of positive scores assigned to each sample across all submitted assays.

[0072] Figure 9: scLIPS protocol validation in DBS three-screen protocol. The figure shows the comparison results obtained from paired serum and DBS samples from the same individuals. This included samples from patients with type 1 diabetes (T1D), celiac disease (CD), and healthy controls (Control).

[0073] DETAILED DESCRIPTION OF THE INVENTION

[0074] Definitions

[0075] In the context of the invention, the term “recombinant antigen molecule” will be understood as a recombinant antigen molecule derived from a self-antigen molecule. A self-antigen may be found in all cell types or be highly specific for a specific cell type in one organ of the body. It may comprise, but is not limited to, proteins, nucleic acids, carbohydrates, lipids, or various combinations of these. In the context of the invention, the recombinant antigen molecules are derived from an antigen molecule expressed by pancreatic beta cells and includes, but is not limited to human insulin, proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, and human Zinc transporter 8. The recombinant antigen molecule according to the invention can be modified by site-directed mutagenesis of coding sequences to change specific amino acid residues or to introduce restriction sites at desired location in the coding sequence or to produce truncated forms. According to the invention, the recombinant antigen molecules comprise theregulatory region of a target gene, which is preferably fused with the DNA coding sequence of the reporter protein.

[0076] In the context of the invention, the term “antibody” or “antibody molecule” as used herein refers to any functional antibody that is capable of specific binding to the antigen of interest, as generally outlined in chapter 7 of Paul, W. E. (Ed.).: Fundamental Immunology 2nd Ed. Raven Press, Ltd., New York 1989. The skilled person understands the term “specific binding” in the context of antigen-antibody of the present invention. The specific binding of an antibody to its antigen(s) excludes non-specific binding, such as for instance non-specific antibody binding to endogenous Fc receptors (FcRs) or non-specific binding due to ionic and / or hydrophobic interactions. The skilled person is able to detect and measure autoantibodies specifically binding to the recombinant antigen molecules of the present invention.

[0077] An antibody or antibody molecule that react with self-molecules, i.e., self-antigens, is referred to as natural antibody or “autoantibody” or “autoantibody molecule”. Autoantibodies can be used as indicators of disease, e.g., their detection can be used to detect a loss of tolerance and inflammation in a subject with an autoimmune disorder. “Islet autoantibody molecules” are well-known in the field and generally understood as autoantibody molecules with specificity to e.g., human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, and human Zinc transporter 8. The islet autoantibodies detected by the recombinant antigen molecules according to the invention include but are not limited to human IAA (Insulin Auto Antibodies), human GADA (Glutamic Acid Decarboxylase Antibodies), human IA-2A (Islet Antigen 2 Antibodies), and human ZnT8A (Zinc Transporter 8 Antibodies).

[0078] In the context of the invention, when the reporter molecule is a reporter protein such as luciferase or a derivate thereof, then the reporter protein will be fused to the recombinant antigen molecule. The term “fused” refers to the genetic or chemical combination of a reporter protein with another protein to form a single, continuous polypeptide chain. This fusion enables the reporter protein, which may provide detectable or measurable properties (such as fluorescence, luminescence, or enzymatic activity), to be expressed and analyzed in conjunction with the target protein, i.e., the antigen molecule. The resulting fused recombinant antigen molecule retains the functional characteristics of both the reporter and the antigen molecule.

[0079] The term “bioluminescent reporter protein” is well known in the art and will be understood by the skilled person in the art. A reporter protein referred herein is well known in the art and include, for instance, a luciferase reporter protein, a nanoluciferase reporter protein, and a NanoLuc™ (NLuc) luciferase reporter protein. According to the invention, a substrate of the reporter protein is added to the assay and the reporter protein catalyzes a reaction that produces light. The amount of light produced can be detected and provides a quantitative measure of the level of the reporterprotein in the sample; the level of the reporter protein in a sample may be used to determine the presence and / or level of an antibody molecule in a sample.

[0080] In the context of the method of the present invention, the term “contacting the sample with the recombinant antigen molecules of the test kit” means contacting the recombinant antigen with the sample from a human subject, under the conditions and for a period of time which is sufficient for the determination of the presence of autoantibodies in the sample from a human subject. As used herein, the term "contact" or "contacting" means bringing together, either directly or indirectly, the recombinant antigen molecules with the sample from a human subject. Contacting may occur, for example, in any number of buffers, salts, solutions, or in cell culture medium. In accordance with the invention, the term "single composition" refers to a formulation containing at least one type of molecule. Specifically, a single composition may include at least one, two, or three recombinant antigen molecules.

[0081] In the context of the invention, a "positive sample" refers to a biological sample obtained from a human subject that, when analysed using the method described herein, exhibits characteristics indicative of an increased risk for developing type 1 diabetes. The sample contains one or more autoantibody molecules associated with type 1 diabetes. It will be understood that the amount of the detected autoantibody molecule(s) in the sample exceeds a predetermined threshold value, as established by comparison with a standard curve for known amounts of said autoantibody molecules. The presence and / or quantity of the autoantibody molecule(s) in the sample, when evaluated in the context of the screening method, suggests an elevated likelihood of the subject developing type 1 diabetes. A sample may be classified as "positive" based on either the mere presence of specific autoantibody molecules or when the quantity of such molecules surpasses a defined concentration threshold, as determined by the standard curve comparison. The classification of a sample as "positive" serves as a key indicator in the overall screening process for assessing an individual's risk of developing type 1 diabetes, and may inform subsequent diagnostic or preventive measures. Detection and measurement of islet autoantibodies may be assessed by comparing the level of autoantibodies in a sample from a subject at risk of developing T1 D, to the level of autoantibodies in a control sample, i.e., a sample from a subject that is not at risk of developing T1D, wherein an increase level of autoantibodies indicates an increased probability of developing T1D. Preferably, the level is statistically significant as assessed by appropriate statistical tests which are known in the art.

[0082] In accordance with the invention, Glutamate Decarboxylase 65 is also known under the following synonyms: GAD65, GAD2, Glutamate Decarboxylase 2, Glutamic Acid Decarboxylase 2, Glutamic Acid Decarboxylase 65, 65 kDa Glutamic Acid Decarboxylase, GAD65 kDa, GAD 65 kDa Isoform.In accordance with the invention, Islet antigen 2 is also known under the following synonyms: IA-2, IA2, ICA512, R-PTP-N, IA-2 / PTP, ICA3, insulinoma-associated tyrosinephosphatase-like protein, islet cell antigen 2, islet cell antigen 512, islet cell autoantigen 3, protein tyrosine phosphatase-like N.

[0083] In accordance with the invention, Zinc transporter 8 is also known under the following synonyms: ZnT8, ZNT8, ZnT-8, ZNT-8, solute carrier family 30 member 8.

[0084] In accordance with the invention, Insulin / proinsulin are also known under the following synonyms:

[0085] Sequences

[0086] Preferred amino acid sequences referred to in the present application can be independently selected from the following sequences. The sequences are represented in an N-terminal to C-terminal order; and they are represented in the one-letter amino acid code.

[0087] The following non-limiting exemplary amino acid sequences were used in the experimental examples of the present application and / or are disclosed herein as preferred amino acid sequences in accordance with the invention: Insulin, Preproinsulin, INS, IDDM, IDDM1, IDDM2, ILPR, IRDN, MODY10, PNDM4.

[0088] Legend of Table 1:

[0089] XXX Nanoluc luciferase reporter sequence (ATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTG GACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAA CTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATC ATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGT GTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACG GGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGA CGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGC CTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGG CGGCTGTGCGAACGCATTCTGGCG (SEQ ID NO: 10))

[0090] XXX Glycine linker

[0091] XXX Signal peptide

[0092] XXX Alanine spacer

[0093] XXX Furin site

[0094] XXX Flexible linker

[0095] XXX “T4 trimer foldon motif’, or “spacer”, or “start” or “stop” codon

[0096] Hl Glycine serine flexible linker

[0097] XXX Antigen sequenceSEQ ID NO: 1 (Insulin B NLuc) ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGA CCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTA CCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAA GATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAG GGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTG TCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTG AGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCA TCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGA TCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGT AACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGG CTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCAT TCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAG CTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGT CCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCT GGAGAACTACTGCAACTAG

[0098] Insulin B NLuc ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 1)

[0099] NLuc IA-2ic ATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAA CCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCC GTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGG TGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATC GACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGT TCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGA GCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGG CTGGCGGCTGTGCGAACGCATTCTGGCGGGTGGAGGCTCTAGAGAATTCACGCGTGGTAC CGCCGCCACCATGGAGCGGCAGCAAGACAAGGAGCGCCTGGCAGCCCTGGGGCCTGAGG GGGCCCATGGTGACACTACCTTTGAGTACCAGGACCTGTGCCGCCAGCACATGGCCACGA AGTCCTTGTTCAACCGGGCAGAGGGTCCACCGGAGCCTTCACGGGTGAGCAGTGTGTCCT CCCAGTTCAGCGACGCAGCCCAGGCCAGCCCCAGCTCCCACAGCAGCACCCCGTCCTGG TGCGAGGAGCCGGCCCAAGCCAACATGGACATCTCCACGGGACACATGATTCTGGCATACA TGGAGGATCACCTGCGGAACCGGGACCGCCTTGCCAAGGAGTGGCAGGCCCTCTGTGCC TACCAAGCAGAGCCAAACACCTGTGCCACCGCGCAGGGGGAGGGCAACATCAAAAAGAAC CGGCATCCTGACTTCCTGCCCTATGACCATGCCCGCATAAAACTGAAGGTGGAGAGCAGCC CTTCTCGGAGCGATTACATCAACGCCAGCCCCATTATTGAGCATGACCCTCGGATGCCAGCC TACATAGCCACGCAGGGCCCGCTGTCCCATACCATCGCAGACTTCTGGCAGATGGTGTGGG AGAGCGGCTGCACCGTCATCGTCATGCTGACCCCGCTGGTGGAGGATGGTGTCAAGCAGT GTGACCGCTACTGGCCAGATGAGGGTGCCTCCCTCTACCACGTATATGAGGTGAACCTGGT GTCGGAGCACATCTGGTGCGAGGACTTCCTGGTGCGGAGCTTCTACCTGAAGAACGTGCA GACCCAGGAGACGCGCACGCTCACGCAGTTCCACTTCCTCAGCTGGCCGGCAGAGGGCA CACCGGCCTCCACGCGGCCCCTGCTGGACTTCCGCAGGAAGGTGAACAAGTGCTACCGG GGCCGCTCCTGCCCCATCATCGTGCACTGCAGTGATGGTGCGGGGAGGACCGGCACCTAC ATCCTCATCGACATGGTCCTGAACCGCATGGCAAAAGGAGTGAAGGAGATTGACATCGCTG CCACCCTGGAGCATGTCCGTGACCAGCGGCCTGGCCTTGTCCGCTCTAAGGACCAGTTTG AGTTTGCCCTGACAGCCGTGGCGGAGGAAGTGAATGCCATCCTCAAGGCCCTGCCCCAGT GA (SEQ ID NO: 2)

[0100] Dual NLuc IA-2ic ATGGAGCGGCAGCAAGACAAGGAGCGCCTGGCAGCCCTGGGGCCTGAGGGGGCC CATGGTGACACTACCTTTGAGTACCAGGACCTGTGCCGCCAGCACATGGCCACGAAGTCCT TGTTCAACCGGGCAGAGGGTCCACCGGAGCCTTCACGGGTGAGCAGTGTGTCCTCCCAGT TCAGCGACGCAGCCCAGACCAGCCCCAGCTCCCACAGCAGCACCCCGTCCTGGTGCGAG GAGCCGGCCCAAGCCAACATGGACATCTCCACGGGACACATGATTCTGGCATACATGGAGG ATCACCTGCGGAACCGGGACCGCCTTGCCAAGGAGTGGCAGGCCCTCTGTGCCTACCAAG CAGAGCCAAACACCTGTGCCACCGCGCAGGGGGAGGGCAACATCAAAAAGAACCGGCATC CTGACTTCCTGCCCTATGACCATGCCCGCATAAAACTGAAGGTGGAGAGCAGCCCTTCTCG GAGCGATTACATCAACGCCAGCCCCATTATTGAGCATGACCCTCGGATGCCAGCCTACATAGCCACGCAGGGCCCGCTGTCCCATACCATCGCAGACTTCTGGCAGATGGTGTGGGAGAGCG GCTGCACCGTCATCGTCATGCTGACCCCGCTGGTGGAGGATGGTGTCAAGCAGTGTGACC GCTACTGGCCAGATGAGGGTGCCTCCCTCTACCACGTATATGAGGTGAACCTGGTGTCGGA GCACATCTGGTGCGAGGACTTCCTGGTGCGGAGCTTCTACCTGAAGAACGTGCAGACCCA GGAGACGCGCACGCTCACGCAGTTCCACTTCCTCAGCTGGCCGGCAGAGGGCACACCGG CCTCCACGCGGCCCCTGCTGGACTTCCGCAGGAAGGTGAACAAGTGCTACCGGGGCCGC TCCTGCCCCATCATCGTGCACTGCAGTGATGGTGCGGGGAGGACCGGCACCTACATCCTCA TCGACATGGTCCTGAACCGCATGGCAAAAGGAGTGAAGGAGATTGACATCGCTGCCACCCT GGAGCATGTCCGTGACCAGCGGCCTGGCCTTGTCCGCTCTAAGGACCAGTTTGAGTTTGC CCTGACAGCCGTGGCGGAGGAAGTGAATGCCATCCTCAAGGCCCTGCCCCAGGGTACCGG CGGGGGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTA CAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTG TCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCC ATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTT AAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGT AATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCC GTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCG ACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGA CCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGGCGCGAGCGGTGGTGGCGGGAGCGG AGGTGGAGGGTCGTCAGGTGCGGCCGCAGAGCGGCAGCAAGACAAGGAGCGCCTGGCA GCCCTGGGGCCTGAGGGGGCCCATGGTGACACTACCTTTGAGTACCAGGACCTGTGCCGC CAGCACATGGCCACGAAGTCCTTGTTCAACCGGGCAGAGGGTCCACCGGAGCCTTCACGG GTGAGCAGTGTGTCCTCCCAGTTCAGCGACGCAGCCCAGACCAGCCCCAGCTCCCACAGC AGCACCCCGTCCTGGTGCGAGGAGCCGGCCCAAGCCAACATGGACATCTCCACGGGACAC ATGATTCTGGCATACATGGAGGATCACCTGCGGAACCGGGACCGCCTTGCCAAGGAGTGG CAGGCCCTCTGTGCCTACCAAGCAGAGCCAAACACCTGTGCCACCGCGCAGGGGGAGGG CAACATCAAAAAGAACCGGCATCCTGACTTCCTGCCCTATGACCATGCCCGCATAAAACTGA AGGTGGAGAGCAGCCCTTCTCGGAGCGATTACATCAACGCCAGCCCCATTATTGAGCATGA CCCTCGGATGCCAGCCTACATAGCCACGCAGGGCCCGCTGTCCCATACCATCGCAGACTTC TGGCAGATGGTGTGGGAGAGCGGCTGCACCGTCATCGTCATGCTGACCCCGCTGGTGGAG GATGGTGTCAAGCAGTGTGACCGCTACTGGCCAGATGAGGGTGCCTCCCTCTACCACGTAT ATGAGGTGAACCTGGTGTCGGAGCACATCTGGTGCGAGGACTTCCTGGTGCGGAGCTTCT ACCTGAAGAACGTGCAGACCCAGGAGACGCGCACGCTCACGCAGTTCCACTTCCTCAGCT GGCCGGCAGAGGGCACACCGGCCTCCACGCGGCCCCTGCTGGACTTCCGCAGGAAGGTG AACAAGTGCTACCGGGGCCGCTCCTGCCCCATCATCGTGCACTGCAGTGATGGTGCGGGG AGGACCGGCACCTACATCCTCATCGACATGGTCCTGAACCGCATGGCAAAAGGAGTGAAGGAGATTGACATCGCTGCCACCCTGGAGCATGTCCGTGACCAGCGGCCTGGCCTTGTCCGCT CTAAGGACCAGTTTGAGTTTGCCCTGACAGCCGTGGCGGAGGAAGTGAATGCCATCCTCAA GGCCCTGCCCCAGTAA (SEQ ID NO: 3)

[0101] NLuc IA-2ic trimeric ATGGAGCGGCAGCAAGACAAGGAGCGCCTGGCAGCCCTGGGGCCTGAGGGGGCC CATGGTGACACTACCTTTGAGTACCAGGACCTGTGCCGCCAGCACATGGCCACGAAGTCCT TGTTCAACCGGGCAGAGGGTCCACCGGAGCCTTCACGGGTGAGCAGTGTGTCCTCCCAGT TCAGCGACGCAGCCCAGGCCAGCCCCAGCTCCCACAGCAGCACCCCGTCCTGGTGCGAG GAGCCGGCCCAAGCCAACATGGACATCTCCACGGGACACATGATTCTGGCATACATGGAGG ATCACCTGCGGAACCGGGACCGCCTTGCCAAGGAGTGGCAGGCCCTCTGTGCCTACCAAG CAGAGCCAAACACCTGTGCCACCGCGCAGGGGGAGGGCAACATCAAAAAGAACCGGCATC CTGACTTCCTGCCCTATGACCATGCCCGCATAAAACTGAAGGTGGAGAGCAGCCCTTCTCG GAGCGATTACATCAACGCCAGCCCCATTATTGAGCATGACCCTCGGATGCCAGCCTACATAG CCACGCAGGGCCCGCTGTCCCATACCATCGCAGACTTCTGGCAGATGGTGTGGGAGAGCG GCTGCACCGTCATCGTCATGCTGACCCCGCTGGTGGAGGATGGTGTCAAGCAGTGTGACC GCTACTGGCCAGATGAGGGTGCCTCCCTCTACCACGTATATGAGGTGAACCTGGTGTCGGA GCACATCTGGTGCGAGGACTTCCTGGTGCGGAGCTTCTACCTGAAGAACGTGCAGACCCA GGAGACGCGCACGCTCACGCAGTTCCACTTCCTCAGCTGGCCGGCAGAGGGCACACCGG CCTCCACGCGGCCCCTGCTGGACTTCCGCAGGAAGGTGAACAAGTGCTACCGGGGCCGC TCCTGCCCCATCATCGTGCACTGCAGTGATGGTGCGGGGAGGACCGGCACCTACATCCTCA TCGACATGGTCCTGAACCGCATGGCAAAAGGAGTGAAGGAGATTGACATCGCTGCCACCCT GGAGCATGTCCGTGACCAGCGGCCTGGCCTTGTCCGCTCTAAGGACCAGTTTGAATTTGCC CTGACAGCCGTGGCGGAGGAAGTGAATGCCATCCTCAAGGCCCTGCCCCAGGGTACCTCT AGAGTCGACCCGGGCGGCCGCGGTGGAGGCATGGTCTTCACACTCGAAGATTTCGTTGGG GACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCC AGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTG AAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAA ATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTG ATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGG ACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTG TGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCC GAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTG CTGCGGGCTACATCCCCGAGGCCCCCAGGGACGGCCAGGCCTACGTGAGGAAGGACGGC GAGTGGGTGCTGCTGAGCACCTTCCTGTAA (SEQ ID NO: 4)

[0102] Dual NLuc ZnT8 dimerATGGAGAAGGACTTCTCCATCTTACTCATGGAAGGTGTGCCAAAGAGCCTGAATTAC AGTGGTGTGAAAGAGCTTATTTTAGCAGTCGACGGGGTGCTGTCTGTGCACAGCCTGCACA TCTGGTCTCTAACAATGAATCAAGTAATTCTCTCAGCTCATGTTGCTACAGCAGCCAGCTGG GACAGCCAAGTGGTTCGGAGAGAAATTGCTAAAGCCCTTAGCAAAAGCTTTACGATGCACTC ACTCACCATTCAGATGGAATCTCCAGTTGACCAGGACCCCGACTGCCTTTTCTGTGAAGACC CCTGTGACGGCGGCGGCAAGGACTTCTCCATCTTACTCATGGAAGGTGTGCCAAAGAGCCT GAATTACAGTGGTGTGAAAGAGCTTATTTTAGCAGTCGACGGGGTGCTGTCTGTGCACAGC CTGCACATCTGGTCTCTAACAATGAATCAAGTAATTCTCTCAGCTCATGTTGCTACAGCAGCC AGCCGGGACAGCCAAGTGGTTCGGAGAGAAATTGCTAAAGCCCTTAGCAAAAGCTTTACGA TGCACTCACTCACCATTCAGATGGAATCTCCAGTTGACCAGGACCCCGACTGCCTTTTCTGT GAAGACCCCTGTGACGgtaccGGCGGGGGTATGGTCTTCACACTCGAAGATTTCGTTGGGGA CTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAG TTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAA ATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGtgcgaacgcattctggcgGGCgcGAGCGGTGGTG GCGGGAGCGGAGGTGGAGGGTCGTCAGGTGCggccGCgGAATTCGCCGCCACCATGGAGA AGGACTTCTCCATCTTACTCATGGAAGGTGTGCCAAAGAGCCTGAATTACAGTGGTGTGAAA GAGCTTATTTTAGCAGTCGACGGGGTGCTGTCTGTGCACAGCCTGCACATCTGGTCTCTAA CAATGAATCAAGTAATTCTCTCAGCTCATGTTGCTACAGCAGCCAGCTGGGACAGCCAAGTG GTTCGGAGAGAAATTGCTAAAGCCCTTAGCAAAAGCTTTACGATGCACTCACTCACCATTCA GATGGAATCTCCAGTTGACCAGGACCCCGACTGCCTTTTCTGTGAAGACCCCTGTGACGGC GGCGGCAAGGACTTCTCCATCTTACTCATGGAAGGTGTGCCAAAGAGCCTGAATTACAGTG GTGTGAAAGAGCTTATTTTAGCAGTCGACGGGGTGCTGTCTGTGCACAGCCTGCACATCTG GTCTCTAACAATGAATCAAGTAATTCTCTCAGCTCATGTTGCTACAGCAGCCAGCCGGGACA GCCAAGTGGTTCGGAGAGAAATTGCTAAAGCCCTTAGCAAAAGCTTTACGATGCACTCACTC ACCATTCAGATGGAATCTCCAGTTGACCAGGACCCCGACTGCCTTTTCTGTGAAGACCCCT GTGACTAG (SEQ ID NO: 5)

[0103] NLuc GAD65 aa96_585 ATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAA CCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCC GTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGT CATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATC GACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGT TCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGA GCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGG CTGGCGGCTGTGCGAACGCATTCTGGCGGGTGGAGGCTCTAGAGAATTCACGCGTGGTAC CGCCGCCACCATGGACCTGCTGCCGGCGTGTGATGGAGAAAGGCCCACTTTGGCGTTTCT GCAAGATGTTATGAACATTTTACTTCAGTATGTGGTGAAAAGTTTCGATAGATCAACCAAAGT GATTGATTTCCATTATCCTAATGAGCTTCTCCAAGAATATAATTGGGAATTGGCAGACCAACCA CAAAATTTGGAGGAAATTTTGATGCATTGCCAAACAACTCTAAAATATGCAATTAAAACAGGG CATCCTAGATACTTCAATCAACTTTCTACTGGTTTGGATATGGTTGGATTAGCAGCAGACTGG CTGACATCAACAGCAAATACTAACATGTTCACCTATGAAATTGCTCCAGTATTTGTGCTTTTGG AATATGTCACACTAAAGAAAATGAGAGAAATCATTGGCTGGCCAGGGGGCTCTGGCGATGG GATATTTTCTCCCGGTGGCGCCATATCTAACATGTATGCCATGATGATCGCACGCTTTAAGAT GTTCCCAGAAGTCAAGGAGAAAGGAATGGCTGCTCTTCCCAGGCTCATTGCCTTCACGTCT GAACATAGTCATTTTTCTCTCAAGAAGGGAGCTGCAGCCTTAGGGATTGGAACAGACAGCGT GATTCTGATTAAATGTGATGAGAGAGGGAAAATGATTCCATCTGATCTTGAAAGAAGGATTCT TGAAGCCAAACAGAAAGGGTTTGTTCCTTTCCTCGTGAGTGCCACAGCTGGAACCACCGTG TACGGAGCATTTGACCCCCTCTTAGCTGTCGCTGACATTTGCAAAAAGTATAAGATCTGGATG CATGTGGATGCAGCTTGGGGTGGGGGATTACTGATGTCCCGAAAACACAAGTGGAAACTGA GTGGCGTGGAGAGGGCCAACTCTGTGACGTGGAATCCACACAAGATGATGGGAGTCCCTT TGCAGTGCTCTGCTCTCCTGGTTAGAGAAGAGGGATTGATGCAGAATTGCAACCAAATGCAT GCCTCCTACCTCTTTCAGCAAGATAAACATTATGACCTGTCCTATGACACTGGAGACAAGGC CTTACAGTGCGGACGCCACGTTGATGTTTTTAAACTATGGCTGATGTGGAGGGCAAAGGGG ACTACCGGGTTTGAAGCGCATGTTGATAAATGTTTGGAGTTGGCAGAGTATTTATACAACATC ATAAAAAACCGAGAAGGATATGAGATGGTGTTTGATGGGAAGCCTCAGCACACAAATGTCTG CTTCTGGTACATTCCTCCAAGCTTGCGTACTCTGGAAGACAATGAAGAGAGAATGAGTCGCC TCTCGAAGGTGGCTCCAGTGATTAAAGCCAGAATGATGGAGTATGGAACCACAATGGTCAG CTACCAACCCTTGGGAGACAAGGTCAATTTCTTCCGCATGGTCATCTCAAACCCAGCGGCA ACTCACCAAGACATTGACTTCCTGATTGAAGAAATAGAACGCCTTGGACAAGATTTATAA

[0104] (SEQ ID NO: 6)

[0105] GAD65aa96-585 competitor ATGGACCTGCTGCCGGCGTGTGATGGAGAAAGGCCCACTTTGGCGTTTCTGCAAGA TGTTATGAACATTTTACTTCAGTATGTGGTGAAAAGTTTCGATAGATCAACCAAAGTGATTGAT TTCCATTATCCTAATGAGCTTCTCCAAGAATATAATTGGGAATTGGCAGACCAACCACAAAATT TGGAGGAAATTTTGATGCATTGCCAAACAACTCTAAAATATGCAATTAAAACAGGGCATCCTA GATACTTCAATCAACTTTCTACTGGTTTGGATATGGTTGGATTAGCAGCAGACTGGCTGACATCAACAGCAAATACTAACATGTTCACCTATGAAATTGCTCCAGTATTTGTGCTTTTGGAATATGT CACACTAAAGAAAATGAGAGAAATCATTGGCTGGCCAGGGGGCTCTGGCGATGGGATATTTT CTCCCGGTGGCGCCATATCTAACATGTATGCCATGATGATCGCACGCTTTAAGATGTTCCCAG AAGTCAAGGAGAAAGGAATGGCTGCTCTTCCCAGGCTCATTGCCTTCACGTCTGAACATAGT CATTTTTCTCTCAAGAAGGGAGCTGCAGCCTTAGGGATTGGAACAGACAGCGTGATTCTGAT TAAATGTGATGAGAGAGGGAAAATGATTCCATCTGATCTTGAAAGAAGGATTCTTGAAGCCAA ACAGAAAGGGTTTGTTCCTTTCCTCGTGAGTGCCACAGCTGGAACCACCGTGTACGGAGCA TTTGACCCCCTCTTAGCTGTCGCTGACATTTGCAAAAAGTATAAGATCTGGATGCATGTGGAT GCAGCTTGGGGTGGGGGATTACTGATGTCCCGAAAACACAAGTGGAAACTGAGTGGCGTG GAGAGGGCCAACTCTGTGACGTGGAATCCACACAAGATGATGGGAGTCCCTTTGCAGTGCT CTGCTCTCCTGGTTAGAGAAGAGGGATTGATGCAGAATTGCAACCAAATGCATGCCTCCTAC CTCTTTCAGCAAGATAAACATTATGACCTGTCCTATGACACTGGAGACAAGGCCTTACAGTG CGGACGCCACGTTGATGTTTTTAAACTATGGCTGATGTGGAGGGCAAAGGGGACTACCGGG TTTGAAGCGCATGTTGATAAATGTTTGGAGTTGGCAGAGTATTTATACAACATCATAAAAAACC GAGAAGGATATGAGATGGTGTTTGATGGGAAGCCTCAGCACACAAATGTCTGCTTCTGGTAC ATTCCTCCAAGCTTGCGTACTCTGGAAGACAATGAAGAGAGAATGAGTCGCCTCTCGAAGG TGGCTCCAGTGATTAAAGCCAGAATGATGGAGTATGGAACCACAATGGTCAGCTACCAACCC TTGGGAGACAAGGTCAATTTCTTCCGCATGGTCATCTCAAACCCAGCGGCAACTCACCAAG ACATTGACTTCCTGATTGAAGAAATAGAACGCCTTGGACAAGATTTATAA (SEQ ID NO: 7) proinsulin competitor ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCTGGTGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAG CATCTGCTCcctctaccagctggagaactactgcaactag (SEQ ID NO: 8)

[0106] hPPI COMPETITOR- CDS ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCTGGTGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAG CATCTGCTCcctctaccagctggagaactactgcaactag (SEQ ID NO: 9)

[0107] Insulin B NLuc Delta B24-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGAACGAGGCATGGTCTTCACACTCGAAGATTTCGTTGGGGACTG GCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTT GTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAAT GGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGG GCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCC TGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGG CCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGA ACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGT AACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTGCTGC GCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGG CGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTGGCATTGTGG AACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAG (SEQ ID NO: 11)

[0108] Insulin B NLuc Delta B25-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCATGGTCTTCACACTCGAAGATTTCGTTGGGGA CTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAG TTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAA ATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTGCTG CGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAG GCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTGGCATTGTG GAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAG (SEQ ID NO: 12)

[0109] Insulin B NLuc Delta B26-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCATGGTCTTCACACTCGAAGATTTCGTTGG GGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTC CAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGT GAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGG TGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTC GGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACC CTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGT TCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTG CTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGG CCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTGGCA TTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAG

[0110] (SEQ ID NO: 13)

[0111] Insulin B NLuc Delta B27-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGG GGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTG GCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAAC TAG (SEQ ID NO: 14)

[0112] Insulin B NLuc Delta B28-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACAATGGTCTTCACACTCGAAGATTTC GTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGT GTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAG CGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGC GACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTT AAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTA TTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGG ACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGG CTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGG GGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGT GGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAA CTAG (SEQ ID NO: 15)

[0113] Insulin B NLuc mut A E17A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGCGAACTACTGCAACTAG (SEQ ID NO: 16)

[0114] Insulin B NLuc mut A L13A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcGCctaccagct ggagaactactgcaactag (SEQ ID NO: 17)

[0115] Insulin B NLuc mut A L13W ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCTGGTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 18)

[0116] Insulin B NLuc mut A Y14A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCGCCCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 19)

[0117] Insulin B NLuc mut B F1AATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCGCTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 20)

[0118] Insulin B NLuc mut B F25D ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCGACTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 21)

[0119] Insulin B NLuc mut B L17A V18A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACGCAGCGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 22)

[0120] Insulin B NLuc mut B L17A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACGCAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 23)

[0121] Insulin B NLuc mut B R22A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAAGCAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 24)

[0122] Insulin B NLuc mut B S9P ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCCCCCACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 25)

[0123] Insulin B NLuc mut B V18A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGCGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 26)

[0124] Insulin B NLuc mut B V2G ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGGGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 27)

[0125] Insulin NLuc A Delta B25-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCCGCCGGAAGCGGGAGGACCTGCAGGTGGGG CAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGAC AGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAAT CTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGA TCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGA AAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGG CACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAA GGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAAC AAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCA ACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTGCTGCGGGCATT GTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAG

[0126] (SEQ ID NO: 28)

[0127] Insulin NLuc A Delta B26-30ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCCGCCGGAAGCGGGAGGACCTGCAGGTG GGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGG CCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACT GGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTT TGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAAT GGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGG GCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCC TGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGG CCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGA ACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGT AACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTGCTGC GGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGC AACTAG (SEQ ID NO: 29)

[0128] Insulin NLucA Delta B27-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACCGCCGGAAGCGGGAGGACCTGCAGG TGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTT GGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGA CTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAG TTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAA ATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTGCTG CGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTG CAACTAG (SEQ ID NO: 30)

[0129] Insulin NLucA Delta B28-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACGCCGGAAGCGGGAGGACCTGC AGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGG GACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCC AGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTG AAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAA ATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTG ATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGG ACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTG TGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCC GAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTG CTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTA CTGCAACTAG (SEQ ID NO: 31)

[0130] Insulin NLucA Delta B30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGCGCCGGAAGCGGGAGG ACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCT GCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGATTT CGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGG TGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGA GCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGG CGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACT TTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGAC TATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAG GGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCT GCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGC GGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTG GAGAACTACTGCAACTAG (SEQ ID NO: 32)

[0131] Insulin NLuc A mut A L13A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCGCCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 33)

[0132] Insulin NLuc A mut A L13W ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCTGGTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 34)

[0133] Insulin NLuc A mut B F1A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCGCTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 35)

[0134] Insulin NLuc A mut B F25D ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCGACTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 36)

[0135] Insulin NLuc A mut B L17A V18A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACGCAGCGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 37)

[0136] Insulin NLuc A mut B L17AATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACGCAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 38)

[0137] Insulin NLuc A mut B R22A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 39)

[0138] Insulin NLuc A mut B V18A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGCGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 40)

[0139] Insulin NLuc A mut B V2G ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGGGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 41)

[0140] Insulin NLuc A mut BR22A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAAGCAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 42)

[0141] Insulin A NLuc ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTG TACCAGCATCTGCTCcctctaccagctggagaactactgcaacGGTGGAGGCATGGTCTTCACACTCGA AGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACA GGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATT GTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCT GAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGAT CATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACAT GATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACT GTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACG GCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCA TTCTGGCGtag (SEQ ID NO: 43)

[0142] Insulin B NLuc ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGAAGCGGGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 44)

[0143] Insulin NLucA ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCGACGCAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGC GGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 45)

[0144] secNLuc insulin ATGAACTCCTTCTCCACAAGCGCCTTCGGTCCAGTTGCCTTCTCCCTGGGCCTGCT CCTGGTGTTGCCTGCTGCCTTCCCTGCCCCAGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGGTGGAGGCT TTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGA ACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGCGGGAGGACCTGCAGGTGGGGC AGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCT GGAGGGGTCCCGACGCAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCT CTACCAGCTGGAGAACTACTGCAACTAG (SEQ ID NO: 46)

[0145] pCMVTnT INS Bchaindel24-30NlucTCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACtcgaGaattCATGGCCCTGTGGAT GCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCCT TTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGA ACGAGGCATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTA CAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTG TCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCC ATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTT AAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGT AATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCC GTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCG ACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGA CCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCGCCGGAAGcggGAGGACCT GCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGGCAGCCTGCAGCCCTTGGC CCTGGAGGGGTCCcgacgcAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcctc taccagctggagaactactgcaactagGaattCACGCGTGgtacCTCTAGAGTCGACCCGGGCGGCCGCTT CGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTAGAATGCAGTGAAA AAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTAACCATTATAAGCTGCAATA AACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGGGGAGATGTGGGAG GTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAAT GGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGA CCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGC CACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTT AGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGC CATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGA CTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGG ATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATT TTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGCGGT ATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGCCAG CCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTGTCTGCTCCCGGCATCC GCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAGAGGTTTTCACCGTCAT CACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTTTATAGGTTAATGTCATG ATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTAT TTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGC TTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTT TTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATG CTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGAT CCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATG TGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATT CTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACA GTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTG ACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAA CTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACAC CACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTC TAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTG CGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGT CTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTAC ACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCT CACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGATTGATTTAAAA CTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCATGACCAAAATCCC TTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGATCTTCTT GAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCG GTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAG AGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACT CTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGT CGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAA CTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCG GACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGG GGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGAT TTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTT ACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT (SEQ ID NO: 47) pCMVTnT INS Bchaindel25-30Nluc TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACtcgaGaattCATGGCCCTGTGGAT GCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCCT TTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGA ACGAGGCTTCATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGG CTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGG GTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACA TCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATT TTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTG GTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCG CCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTAT CGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCGCCGGAAGcggGAGGA CCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGGCAGCCTGCAGCCCTT GGCCCTGGAGGGGTCCcgacgcAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCT CcctctaccagctggagaactactgcaactagGaattCACGCGTGgtacCTCTAGAGTCGACCCGGGCGGCC GCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTAGAATGCAGT GAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTAACCATTATAAGCTGC AATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGGGGAGATGTGG GAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTG GCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGG CGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAG CGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTT CTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCC GATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGT GGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAG TGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAA GGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCG AATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGC GGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGC CAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTGTCTGCTCCCGGC ATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAGAGGTTTTCACCG TCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTTTATAGGTTAATGT CATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCC CTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAA TGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCC CTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAG ATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAA GATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGC TATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACA CTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCAT GACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTAC TTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCA TGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGT GACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACT TACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCAC TTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCG TGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAG GTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGATTGA TTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCATGACCA AAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGA TCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTA CCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTT CAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCA AGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCC AGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGC AGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTAC ACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAA AGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTT CCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGC GTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGG CCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT (SEQ I D NO: 48)

[0146] pCMVTnT INS Bchaindel26-30Nluc TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACtcgaGaattCATGGCCCTGTGGAT GCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGA ACGAGGCTTCTTCATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCC GGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCG GGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGA CATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAA TTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACAC TGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATC GCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTA TCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGT GACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCGCCGGAAGcggGAGGA CCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGGCAGCCTGCAGCCCTT GGCCCTGGAGGGGTCCcgacgcAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCT CcctctaccagctggagaactactgcaactagGaattCACGCGTGgtacCTCTAGAGTCGACCCGGGCGGCC GCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTAGAATGCAGT GAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTAACCATTATAAGCTGC AATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGGGGAGATGTGG GAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTG GCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGG CGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAG CGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTT CTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCC GATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGT GGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAG TGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAA GGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCG AATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGC GGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGC CAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTGTCTGCTCCCGGC ATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAGAGGTTTTCACCG TCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTTTATAGGTTAATGT CATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCC CTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAA TGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCC CTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAG ATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAA GATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACA CTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCAT GACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTAC TTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCA TGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGT GACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACT TACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCAC TTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCG TGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTT ATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAGATAG GTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGATTGA TTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCATGACCA AAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGA TCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTA CCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTT CAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCA AGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCC AGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGC AGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTAC ACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAA AGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTT CCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGC GTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGG CCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT (SEQ I D NO: 49)

[0147] pCMVTnT INS Bchaindel27-30Nluc TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAAATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACtcgaGaattCATGGCCCTGTGGAT GCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCCT TTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGA ACGAGGCTTCTTCTACATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACA GCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATC TCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGAT CGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAA AAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGC ACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAG GCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACA AAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAA CGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCGCCGGAAGcg gGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCcgacgcAAGCGTGGCATTGTGGAACAATGCTGTACCAGCA TCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGgtacCTCTAGAGTCGACCCGGG CGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTAGAAT GCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTAACCATTATAA GCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGGGGAGA TGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGG GCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGA ATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCG CAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCC TTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGT TCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGT AGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAA TAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTT ATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAAC GCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCATCT GTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTGTCTGCTCC CGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAGAGGTTTTC ACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTTTATAGGTTA ATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCGGA ACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTG ATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTT ATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTA AAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCG GTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTC TGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCAT ACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGATG GCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCAAC TTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGGGG ATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGA GCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCGAA CTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGG ACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGT GAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTATCG TAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTGAG ATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTAGA TTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCATG ACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAA AGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACC GCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTG GCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCAC TTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGC TGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATAAG GCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACGAC CTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGG AGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGA GCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTT GAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAAC GCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT

[0148] (SEQ ID NO: 50)

[0149] pCMVTnT INS Bchaindel28-30NlucTCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACtcgaGaattCATGGCCCTGTGGAT GCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCCT TTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGA ACGAGGCTTCTTCTACACAATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAG ACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGA ATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAA GATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATC GAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTAT GGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATG AAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCA ACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCAT CAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCGCCGGAA GcggGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGGCAGCCTG CAGCCCTTGGCCCTGGAGGGGTCCcgacgcAAGCGTGGCATTGTGGAACAATGCTGTACCAG CATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGgtacCTCTAGAGTCGACCCG GGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTAGA ATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTAACCATTAT AAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGGGGA GATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGCCT GAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACG CGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTT CCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGG GTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCAC GTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTT AATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGAT TTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTA ACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCAT CTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCATA GTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTGTCTGCT CCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAGAGGTTT TCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTTTATAGGT TAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCG GAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCC TGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCC TTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAG TAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAG CGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAG TTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCG CATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGA TGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCA ACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGG GGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGAC GAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCG AACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCA GGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCG GTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTAT CGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTG AGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTA GATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCA TGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATC AAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCA CCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAAC TGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACC ACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATA AGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACG ACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAG GGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGG GAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGAC TTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAA CGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT

[0150] (SEQ ID NO: 51)

[0151] pCMVTnT insulin Nluc-Achain A L13A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGGC AGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGA GGGGTCCcgacgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGA CAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAA TCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAG ATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCG AAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGA AGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAA CAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATC AACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGTG GAACAATGCTGTACCAGCATCTGCTCcGCctaccagctggagaactactgcaactagGaattCACGCGTGG TACCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTT TGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATT GCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTAT GTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTG GTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTC CCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCG GCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCT CCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAA TCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTT GATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGAC GTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTAT CTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGG TATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCT GCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCT GACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCT GCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGAT ACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTT CGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGC TCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCC AGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATC GAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAAT GATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAG AGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAG TGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCT TTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGA AGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGC AAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAG GCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGG TAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTT ACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCC TTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACC CCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTT TTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCC GTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCC TGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCC ACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTT CGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGA AAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACAT GGCTCGACAGATCT (SEQ ID NO: 52)

[0152] pCMVTnT insulin Nluc-Achain B chain F1A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC GCTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGG GAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGG CAGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGG AGGGGTCCcgacgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAG ACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGA ATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAA GATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATC GAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTAT GGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATG AAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCA ACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCAT CAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGT GGAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGG TACCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTT TGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATT GCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTAT GTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTG GTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTC CCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCG GCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCT CCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAA TCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTT GATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGAC GTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTAT CTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGG TATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCT GCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCT GACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCT GCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGAT ACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTT CGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGC TCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCC AGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAAT GATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAG AGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAG TGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCT TTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGA AGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGC AAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAG GCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTG ATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGG TAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTT ACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCC TTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACC CCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTT TTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCC GTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCC TGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCC ACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTT CGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGA AAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACAT GGCTCGACAGATCT (SEQ ID NO: 53)

[0153] pCMVTnT insulin Nluc-Achain B L17A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGACTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACGCAGTGTGCGGG GAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGG CAGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGG AGGGGTCCcgacgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAG ACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGA ATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAA GATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATC GAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTAT GGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATG AAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCA ACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCAT CAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGT GGAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGG TACCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTT TGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATT GCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTAT GTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTG GTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTC CCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCG GCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCT CCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAA TCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTT GATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGAC GTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTAT CTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCT GCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCT GACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCT GCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGAT ACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTT CGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGC TCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCC AGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATC GAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAAT GATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAG AGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAG TGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCT TTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGA AGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGC AAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAG GCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTG ATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGG TAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTT ACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCC TTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACC CCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTT TTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCC GTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCC TGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCC ACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTT CGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGA AAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACAT GGCTCGACAGATCT (SEQ ID NO: 54)

[0154] pCMVTnT insulin Nluc-Achain B R22ATCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGGC AGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGA GGGGTCCcgacgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGA CAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAA TCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAG ATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCG AAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATG GCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGA AGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAA CAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATC AACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGTG GAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGT ACCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTT GGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATT GCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTAT GTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTC CCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCG GCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCT CCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAA TCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTT GATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGAC GTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTAT CTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGG TATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCT GCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCT GACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCT GCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGAT ACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTT CGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGC TCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCC AGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATC GAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAAT GATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAG AGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAG TGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCT TTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGA AGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGC AAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAG GCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTG ATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGG TAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTT ACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCC TTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACC CCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTT TTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCC GTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCC ACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTT CGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGA AAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACAT GGCTCGACAGATCT (SEQ ID NO: 55)

[0155] pCMVTnT insulin Nluc-Achain Bchain V2G TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGGGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGG GAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGG CAGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGG AGGGGTCCcgacgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAG ACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGA ATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAA GATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATC GAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATG AAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCA ACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCAT CAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGT GGAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGG TACCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTT TGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATT GCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTAT GTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTG GTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTC CCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCG GCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCT CCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAA TCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTT GATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGAC GTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTAT CTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGG TATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCT GCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCT GACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCT GCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGAT ACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTT CGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGC TCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCC AGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATC GAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAAT GATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAG AGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAG TGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCT TTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGA AGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGC AAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAG GCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGG TAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTT ACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCC TTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACC CCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTT TTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCC GTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCC TGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCC ACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTT CGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGA AAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACAT GGCTCGACAGATCT (SEQ ID NO: 56)

[0156] pCMVTnT insulin Nluc-Achain del B26-30 TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCCGCCGGAAGcggGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGC GGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcgacgcAAG CGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACC TGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGT AACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTC ATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGT GGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCG ACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTT CGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAG CGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGC TGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGTGGAACAATGCTGTACCA GCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTAGAGTCGACC CGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTA GAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTAACCATT ATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGGG GAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGAT CCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAGC CTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTA CGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCC CTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTA GGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTC ACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTC TTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTT GATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAAT TTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGC ATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCAT AGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTGTCTGC TCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAGAGGTT TTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTTTATAGG TTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCG GAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCC TGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCC TTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAG TAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAG TTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCG CATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGA TGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCA ACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGG GGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGAC GAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCG AACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCA GGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCG GTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTAT CGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTG AGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTA GATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCA TGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATC AAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCA CCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAAC TGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACC ACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCT GCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATA AGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACG ACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAG GGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGG GAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGAC TTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAA CGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT

[0157] (SEQ ID NO: 57)

[0158] pCMVTnT insulin Nluc-Achain del B27-30 TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGACTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACCGCCGGAAGcggGAGGACCTGCAGGTGGGGCAGGTGGAGCTGG GCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcgacgcA AGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAA CCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCC GTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGT CATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGG TGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATC GACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGT TCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGA GCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGG CTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGTGGAACAATGCTGTACC AGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTAGAGTCGAC CCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAAC TAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTAACCA TTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGG GGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGA TCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGCGCAG CCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTT ACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCC CTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTA GGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTC ACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTC TTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTT GATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAAT TTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCAT AGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTGTCTGC TCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAGAGGTT TTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTTTATAGG TTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGTGCGCG GAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCC TGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGTCGCCC TTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAG TAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAG CGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTTTAAAG TTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTCGCCG CATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTACGGA TGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCGGCCA ACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACATGGG GGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGAC GAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTGGCG AACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCA GGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCG GTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCCGTAT CGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATCGCTG AGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATACTTTA GATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAATCTCA TGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAAGATC AAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCA CCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAAC TGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACC ACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCT GCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACCGGATA AGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCGAACG ACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCCCGAAG GGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCACGAGG GAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCTCTGAC TTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAA CGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT

[0159] (SEQ ID NO: 58)

[0160] pCMVTnT insulin Nluc-Achain del B28-30TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACGCCGGAAGcggGAGGACCTGCAGGTGGGGCAGGTGGAGC TGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcga cgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTA CAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTG TCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCC ATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTT AAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGT AATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCC GTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCG ACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGA CCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGTGGAACAATGCTG TACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTAGAGT CGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCA CAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTTGTA ACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGTTCA GGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTTGC GCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGT GGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTC TTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCC CTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGAT GGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCA CGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATT CTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACA AAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCT TACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGATGC CGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGCTTG TCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGTCAG AGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTATTTT TATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAATGT GCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAAT AACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCCGTGT CGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGCTGGT GAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTC AACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCACTTT TAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTCGGTC GCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCATCTTA CGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACACTGCG GCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCACAACAT GGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAAC GACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATTAACTG GCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTT GCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAG CCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCCTCCC GTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACAGATC GCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATATATA CTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGATAAT CTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAGAAAA GATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAA ACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAG GTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGG CCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACC GGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGC GAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCC CGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCA CGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCT CTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCC AGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTCGAC AGATCT (SEQ ID NO: 59)

[0161] pCMVTnT insulin Nluc-Achain del B30 TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGCGCCGGAAGcggGAGGACCTGCAGGTGGGGCAGG TGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGG GTCCcgacgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAG CCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCT CGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATC GACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAA AAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGAAGG CATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAA ATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACG GAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGTGGAAC AATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCT CTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGA CAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTT TATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTT CAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAA AATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAA CAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGG GTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTT TCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGG GGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTA GGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTG GAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTC GGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTG ATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTAT TTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGC TCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGA CGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGC ATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATAC GCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCG GGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTC ATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAAC ATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAG AAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGA ACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGA TGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAG CAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGA AAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTG ATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTT TTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAA GCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCA AACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGG CGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGT AAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAA TAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTA CTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCT TTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCC CGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCA AACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTT TTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCG TAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCT GTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGA TAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGC TTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCA CGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGA GAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTC GCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAA AAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATG GCTCGACAGATCT (SEQ ID NO: 60)

[0162] pCMVTnT_hPPI furin-Nlucstop TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGGC AGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGA GGGGTCCcgacgcAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcctctaccagctg gagaactactgcaacGGTGGAGGCATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACA GACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAG AATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGA AGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGAT CGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTA TGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTAT GAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGC AACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCA TCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGtagGaattCACGCGTGGT ACCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTT GGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATT GCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTAT GTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTG GTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTC CCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCG GCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCT CCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAA TCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTT GATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGAC GTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTAT CTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGG TATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCT GCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCT GACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCT GCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGAT ACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTT CGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGC TCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCC AGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAAT GATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAG AGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAG TGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCT TTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGA AGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGC AAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAG GCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTG ATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGG TAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTT ACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCC TTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACC CCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTT TTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCC GTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCC TGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCC ACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTT CGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGA AAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACAT GGCTCGACAGATCT (SEQ ID NO: 61)

[0163] pCMVTnT_hPreProlns B chain NIuc furin TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGACTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCG CCGGAAGcggGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggccCTGGT GCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcgacgcAAGCGTGGCATTGTGGAAC AATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCT CTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGA CAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTT TATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTT CAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAA AATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAA CAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGG GTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTT TCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGG GGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTA GGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTG GAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTC GGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTG ATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGC TCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGA CGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGC ATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATAC GCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCG GGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTC ATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAAC ATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAG AAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGA ACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGA TGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAG CAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGA AAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTG ATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTT TTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAA GCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCA AACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGG CGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGA TAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGT AAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAA TAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTA CTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCT TTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCC CGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCA AACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTT TTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCG TAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCT GTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGA TAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGC TTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCA CGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGA GAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTC GCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAA AAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATG GCTCGACAGATCT (SEQ ID NO: 62)

[0164] pCMVTnT_hPreProlns furin Nluc-AchainTCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGGC AGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGA GGGGTCCcgacgcAAGCGTATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGA CAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAA TCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAG ATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCG AAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATG GCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGA AGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAA CAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATC AACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGGGCATTGTG GAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGT ACCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTT GGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATT GCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTAT GTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTC CCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCG GCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCT CCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAA TCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTT GATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGAC GTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTAT CTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAG CTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGG TATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCT GCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCT GACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCT GCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGAT ACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTT CGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGC TCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCA ACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCC AGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATC GAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAAT GATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAG AGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACA GAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAG TGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCT TTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGA AGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGC AAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAG GCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTG ATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGG TAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAA ATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTT ACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCC TTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACC CCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGC AAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTT TTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCC GTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACG ATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAG CTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCC ACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGG AGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTT CGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGA AAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACAT GGCTCGACAGATCT (SEQ ID NO: 63)

[0165] pCMVTnT_sNluc_hPPI_furin_c-peptide TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACtcgaGgaattcGCCGCCACCATGAA CTCCTTCTCCACAAGCGCCTTCGGTCCAGTTGCCTTCTCCCTGGGCCTGCTCCTGGTGTTG CCTGCTGCCTTCCCTGCCCCAGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGA CAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAA TCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAG ATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCG AAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATG GCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGA AGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAA CAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGGTGGAGGCTTTGTGAA CCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGAACGAGG CTTCTTCTACACACCCAAGACCCGCCGGAAGcggGAGGACCTGCAGGTGGGGCAGGTGGA GCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCC cgacgcAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactact gcaactagGAATTCcggtacCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGA TACATTGATGAGTTTGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAA ATTTGTGATGCTATTGCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAA TTGCATTCATTTTATGTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAA CCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGC ACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCG GCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCG CCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCC CCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCG ACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTT TTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAAC AACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTAT TGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTAC AATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGC ACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACAC CCGCTGACGCGCCCTGACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGA CCGTCTCCGGGAGCTGCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGAC GAAAGGGCCTCGTGATACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGAC GTCAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACA TTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGG AAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTT CCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTG CACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCC CGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCG TATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTG AGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGT GCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACC GAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGG AACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAAT GGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAAT TAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCA GCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGG CAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGG TAACTGTCAGACCAAGTTTACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAA AGGATCTAGGTGAAGATCCTTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCG TTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCT GCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCG GATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAA TACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTA CATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTT ACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGG GGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGC GTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAA GCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTAT CTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTC AGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTT TTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT (SEQ ID NO: 64)

[0166] Proinsulin B NLuc Delta B24-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCATGGTCTTCACACTCGAAGATTTCGTTGGGGACTG GCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTT GTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAAT GGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGG GCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCC TGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGG CCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGA ACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGT AACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTGCTGC GCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGG CGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTGGCATTGTGG AACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAG (SEQ ID NO: 65)

[0167] Proinsulin B NLuc Delta B25-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGAACGAGGCTTCATGGTCTTCACACTCGAAGATTTCGTTGGGGA CTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAG TTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAA ATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTGCTGCTG CGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAG GCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTGGCATTGTG GAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAG (SEQ ID NO: 66)

[0168] Proinsulin B NLuc Delta B26-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCATGGTCTTCACACTCGAAGATTTCGTTGG GGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTC CAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGT GAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACC AAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGG TGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTC GGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACC CTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGT TCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGCTG CTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGG CCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTGGCA TTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACTAG

[0169] (SEQ ID NO: 67)

[0170] Proinsulin B NLuc Delta B27-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGG GGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTG GCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAAC TAG (SEQ ID NO: 68)

[0171] Proinsulin B NLuc Delta B28-30 ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACAATGGTCTTCACACTCGAAGATTTC GTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGT GTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAG CGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGC GACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTT AAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTA TTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGG ACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGC TGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGG CTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGG GGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGT GGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAA CTAG (SEQ ID NO: 69)

[0172] Proinsulin B NLuc mut A L13A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCC TGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcGCctaccagctggagaacta ctgcaactag (SEQ ID NO: 70)

[0173] Proinsulin B NLuc mut A L13W ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCC TGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCTGGTACCAGCTGGA GAACTACTGCAACTAG (SEQ ID NO: 71)

[0174] Proinsulin B NLuc mut A Y14A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCGCCCAGCTGGA GAACTACTGCAACTAG (SEQ ID NO: 72)

[0175] Proinsulin B NLuc mut B F1A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCGCTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCC TGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGA GAACTACTGCAACTAG (SEQ ID NO: 73)

[0176] Proinsulin B NLuc mut B L17A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACGCAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCC TGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGA GAACTACTGCAACTAG (SEQ ID NO: 74)

[0177] Proinsulin A NLucATGGCCCTGTGGATGCGCTTGTTGCCCCTTCTGGCCCTTCTCGCACTGTGGGGACC TGATCCAGCTGCGGCCTTTGTGAACCAGCACCTGTGTGGGTCCCATCTGGTGGAAGCCCT GTATCTCGTCTGTGGAGAACGCGGCTTCTTCTACACACCGAAGACCCGCCGGGAGGCAGA GGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAG CCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTGGCATTGTGGAGCAGTGCTGCACCAG CATCTGCTCCCTCTACCAGCTGGAGAACTACTGCAACGAATTCGGTGGAGGCATGGTCTTC ACACTCGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTC CTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCC AAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTAT GAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTG TGGATGATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACG CCGAACATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAA AGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAA CCCCGACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTG CGAACGCATTCTGGCGTAA (SEQ ID NO: 75)

[0178] Proinsulin B NLuc no PG duplication ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGA ACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCC TGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGA GAACTACTGCAACTAG (SEQ ID NO: 76)

[0179] Proinsulin B NLuc ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACT CGAAGATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGG ATTGTCCTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGG TCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGAT GATCATCACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAA CATGATCGACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATC ACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCG ACGGCTCCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAAC GCATTCTGGCGGCTGCTGCTGCGCGCCGGGAGGCAGAGGACCTGCAGGTGGGGCAGGTG GAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGG GGTCCCTGCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCA GCTGGAGAACTACTGCAACTAG (SEQ ID NO: 77)

[0180] Proinsulin C pep NLuc C pep ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaATGGTCTTCACACTCGAAG ATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGG GAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTC CTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAG CGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATC ACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATC GACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAA CAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCT CCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTC TGGCGgctgctgctgcGggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCT GCAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactg caactag (SEQ ID NO: 78)

[0181] Proinsulin NLuc A c peptide PG duplication ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCCCTGGTGCAGGCAG CCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGA TTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGG AGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCC TGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCA CTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCG ACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACA GGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCC CTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTG GCGGCTGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGC TGGAGAACTACTGCAACTAG (SEQ ID NO: 79)

[0182] Proinsulin NLucA ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAG AACTACTGCAACTAG (SEQ ID NO: 80)

[0183] secNLuc Proinsulin ATGAACTCCTTCTCCACAAGCGCCTTCGGTCCAGTTGCCTTCTCCCTGGGCCTGCT CCTGGTGTTGCCTGCTGCCTTCCCTGCCCCAGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGGTGGAGGCT TTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGA ACGAGGCTTCTTCTACACACCCAAGACCCGCCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAG GGGTCCcTgcAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcctctaccagctg gagaactactgcaactag (SEQ ID NO: 81)

[0184] Proinsulin NLuc A mut A L13A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCGCCTACCAGCTGGAG AACTACTGCAACTAG (SEQ ID NO: 82)

[0185] Proinsulin NLuc A mut A L13W ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCTGGTACCAGCTGGAG AACTACTGCAACTAG (SEQ ID NO: 83)

[0186] Proinsulin NLuc A mut A Y14AATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCGCCCAGCTGGAG AACTACTGCAACTAG (SEQ ID NO: 84)

[0187] Proinsulin NLuc A mut B F1A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCGCTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACCTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAG AACTACTGCAACTAG (SEQ ID NO: 85)

[0188] Proinsulin NLuc A mut B L17A V18A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACGCAGCGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAG AACTACTGCAACTAG (SEQ ID NO: 86)

[0189] Proinsulin NLuc A mut B L17A ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGAC CTGACCCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTC TCTACGCAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCCGCCGGGAGGCAG AGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGGGCCCAGGCGCAGGCAGCCTGCA GCCCTTGGCCCTGGAGGGGTCCCTGCAGAAGCGTATGGTCTTCACACTCGAAGATTTCGTT GGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGT GTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGC GGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCG ACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTA AGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTAT TTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGA CCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCT GTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGC TGCTGCTGCGGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCCCTCTACCAGCTGGAG AACTACTGCAACTAG (SEQ ID NO: 87)

[0190] Name Sequence (SEQ ID NO)

[0191] pCMVTnT ProIns B NIucA chain L13A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGGCATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCG CCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGG CAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACAATGC TGTACCAGCATCTGCTCcGCctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTAG AGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAA CCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATT TGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGG TTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATC GATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGT TGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGT GGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGC TTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGG CTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGG TGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAG TCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTA ACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCT CCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGA TGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGC TTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGT CAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTAT TTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAA TGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGA CAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCC GTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGC TGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGA TCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCA CTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTC GGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCA TCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACAC TGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCAC AACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATAC CAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATT AACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATA AAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCT GGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCC TCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACA GATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATA TATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGA TAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAG AAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAA AAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCG AAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTT AGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTAC CAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTT ACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGG AGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCT TCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGC GCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCA CCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTC GACAGATCT (SEQ ID NO: 88)

[0192] pCMVTnT ProIns B NIucA chain L13W TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCG CCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGG CAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACAATGC TGTACCAGCATCTGCTCcTGGtaccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTAG AGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATT TGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGG TTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATC GATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGT TGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGT GGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGC TTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGG CTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGG TGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAG TCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTC TATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTA ACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCT CCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGA TGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGC TTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGT CAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTAT TTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAA TGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGA CAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCC GTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGC TGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGA TCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCA CTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTC GGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCA TCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACAC TGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCAC AACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATAC CAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATT AACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATA AAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCT GGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCC TCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACA GATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATA TATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGA TAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAG AAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCG AAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTT AGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTAC CAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTT ACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGG AGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCT TCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGC GCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCA CCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAAC GCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTC GACAGATCT (SEQ ID NO: 89)

[0193] pCMVTnT Proinsulin B NIucA chain E17A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCG CCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGG CAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACAATGC TGTACCAGCATCTGCTCcctctaccagctggCgaactactgcaactagGaattCACGCGTGGTACCTCTAGA GTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAAC CACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTT GTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGT TCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCG ATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTT GCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGT GGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGC TTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGG CTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGG TGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAG TCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTC TATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTA ACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCT CCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGA TGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGC TTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGT CAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTAT TTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAA TGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGA CAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCC GTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGC TGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGA TCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCA CTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTC GGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCA TCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACAC TGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCAC AACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATT AACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATA AAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCT GGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCC TCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACA GATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATA TATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGA TAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAG AAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAA AAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCG AAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTT AGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTAC CAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTT ACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGG AGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCT TCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGC GCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCA CCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAAC GCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTC GACAGATCT (SEQ ID NO: 90)

[0194] pCMVTnT Proinsulin B NIuc A chain Y14A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGTTTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATG GGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCG CCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGG CAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACAATGC TGTACCAGCATCTGCTCcctcGCccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTAG AGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAA CCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATT TGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGG TTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATC GATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGT TGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGT GGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGC TTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGG CTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGG TGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAG TCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTC TATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTA ACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCT CCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGA TGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGC TTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGT CAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTAT TTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAA TGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCC GTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGC TGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGA TCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCA CTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTC GGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCA TCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACAC TGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCAC AACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATAC CAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATT AACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATA AAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCT GGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCC TCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACA GATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATA TATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGA TAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAG AAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAA AAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCG AAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTT AGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTAC CAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTT ACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGG AGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCT TCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGC GCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCA CCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAAC GCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTC GACAGATCT (SEQ ID NO: 91)

[0195] pCMVTnT Proinsulin B NIuc B chain F1A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATCAAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC GCTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGG GAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGG ACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCA GTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGA AAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAA TGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTG ATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGG ACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTG TGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCC GAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgc GCGCCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGC AGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACA ATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTC TAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGAC AAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTT ATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTC AGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAA ATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAAC AGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGG TGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTT CGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGG GGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAG GGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCG GTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGA TTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATT TTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCT CTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGAC GGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCA TGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACG CCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGG GGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCAT GAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACAT TTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAA ACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAAC TGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATG AGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCA ACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAA AGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATA ACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTT GCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCC ATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAAC TATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCG GATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAA ATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAA GCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATA GACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTAC TCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTT TTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCC GTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAA ACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTT TCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGT AGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTG TTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGAT AGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGC TTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCA CGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGA GAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTC GCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATG GCTCGACAGATCT (SEQ ID NO: 92)

[0196] pCMVTnT Proinsulin B NIuc B chain L17A TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACGCAGTGTGCGGG GAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGG ACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCA GTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGA AAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAA TGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTG ATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGG ACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTG TGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCC GAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgc GCGCCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGC AGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACA ATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTC TAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTT ATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTC AGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAA ATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAAC AGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGG TGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTT CGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGG GGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAG GGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGG AGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCG GTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGA TTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATT TTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCT CTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGAC GGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCA TGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACG CCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGG GGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCAT GAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACAT TTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAA ACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAAC TGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATG AGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCA ACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAA AGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATA ACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTT GCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCC ATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAAC TATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCG GATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAA ATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAA GCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATA GACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTAC TCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTT TTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCC GTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTT TCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGT AGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTG TTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGAT AGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGC TTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCA CGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGA GAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTC GCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAA AAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATG GCTCGACAGATCT (SEQ ID NO: 93)

[0197] pCMVTnT Proinsulin B NIuc no cpeptide PG duplication TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGA TGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCC TTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGG AACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGGAC TGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGT TTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAA TGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATC CTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACG GCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGG AACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAG TAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCG CCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggcGCAGG CAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACAATGC TGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTAGA GTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAAAC CACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTATTT GTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAGGT TCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAATCG ATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACAGTT GCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGT GGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGC TTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGG CTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGG TGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAG TCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTC TATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTA ACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATTTTCT CCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCTCTGA TGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGACGGGC TTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCATGTGT CAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACGCCTAT TTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGGGGAAA TGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGA CAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACATTTCC GTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAAACGC TGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAACTGGA TCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATGAGCA CTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCAACTC GGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAAAGCA TCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATAACAC TGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTTGCAC AACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAACTATT AACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATA AAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAAATCT GGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAAGCCC TCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATAGACA GATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTACTCATA TATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTTTTTGA TAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCCGTAG AAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAA AAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCG AAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTT AGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTAC CAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTT ACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGG AGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCT TCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGC GCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCA CCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAAC GCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGGCTC GACAGATCT (SEQ ID NO: 94)

[0198] pCMVTnT_hPreProlns B chain Nlucpeptide TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTATTGGCC ATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGACCGCC ATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTCATAGCC CATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCGCCCAAC GACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGGGACTTT CCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATCAAGTGT ATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGGCATTAT GCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTCATCGCT ATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGACTCACG GGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAAATCAAC GGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAGGCGTGT ACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAAGCTTTA TTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAGTCTCGA ACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTACAAGACA GGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGTTTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCACTCCCAG TTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTTAGGTGA CACTATAGAATACAAGCTACTTGTTCTTTTTGCACTCGAGaattCATGGCCCTGTGGATGCGCC TCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGACCCAGCCGCAGCCTTTGTGA ACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGAACGAG GCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCG ACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTT CAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGC TGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCA GATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCA CTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCG TATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACG GCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAAC CATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGgctgctgctgcGCGCCG GgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGCGGGggccCaggccCTGGTGCA GGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAAGCGTGGCATTGTGGAACAAT GCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactagGaattCACGCGTGGTACCTCTA GAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAGATACATTGATGAGTTTGGACAA ACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGATGCTATTGCTTTAT TTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACAATTGCATTCATTTTATGTTTCAG GTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAAACCTCTACAAATGTGGTAAAAT CGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCGCACCGATCGCCCTTCCCAACA GTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGT GTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTC GCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGG GGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAG GGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGG AGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCG GTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGA TTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTACAATTTCCTGATGCGGTATT TTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTGCACTCTCAGTACAATCTGCT CTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACACCCGCTGACGCGCCCTGAC GGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTGACCGTCTCCGGGAGCTGCA TGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGACGAAAGGGCCTCGTGATACG CCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGTCAGGTGGCACTTTTCGG GGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATACATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGGAAGAGTATGAGTATTCAACAT TTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTTCCTGTTTTTGCTCACCCAGAA ACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTGCACGAGTGGGTTACATCGAAC TGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCCCGAAGAACGTTTTCCAATGATG AGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCGTATTGACGCCGGGCAAGAGCA ACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTGAGTACTCACCAGTCACAGAAA AGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGTGCTGCCATAACCATGAGTGATA ACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACCGAAGGAGCTAACCGCTTTTTT GCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGGAACCGGAGCTGAATGAAGCC ATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAATGGCAACAACGTTGCGCAAAC TATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCG GATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGCTGGCTGGTTTATTGCTGATAA ATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCAGCACTGGGGCCAGATGGTAA GCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGGCAACTATGGATGAACGAAATA GACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGGTAACTGTCAGACCAAGTTTAC TCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAAAGGATCTAGGTGAAGATCCTT TTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCGTTCCACTGAGCGTCAGACCCC GTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAA ACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTT TCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGT AGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTG TTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGAT AGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGC TTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCA CGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGA GAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTC GCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAA AAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATG GCTCGACAGATCT (SEQ ID NO: 95)

[0199] PreProins B chain NIuc c peptide PG duplication ATGGCCCTGTGGATGCGCCTCCTGCCCCTGCTGGCGCTGCTGGCCCTCTGGGGACCTGAC CCAGCCGCAGCCTTTGTGAACCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTAC CTAGTGTGCGGGGAACGAGGCTTCTTCTACACACCCAAGACCATGGTCTTCACACTCGAAG ATTTCGTTGGGGACTGGCGACAGACAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGG GAGGTGTGTCCAGTTTGTTTCAGAATCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTC CTGAGCGGTGAAAATGGGCTGAAGATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCGAAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATC ACTTTAAGGTGATCCTGCACTATGGCACACTGGTAATCGACGGGGTTACGCCGAACATGATC GACTATTTCGGACGGCCGTATGAAGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAA CAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCT CCCTGCTGTTCCGAGTAACCATCAACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTC TGGCGgctgctgctgcGCGCCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGAGCTGGGC GGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCCcTgcAGAA GCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactactgcaactag (SEQ ID NO: 96)

[0200] pCMVT nT_sNluc_hPPI_c-peptide TCAATATTGGCCATTAGCCATATTATTCATTGGTTATATAGCATAAATCAATATTGGCTAT TGGCCATTGCATACGTTGTATCTATATCATAATATGTACATTTATATTGGCTCATGTCCAATATGA CCGCCATGTTGGCATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTC ATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCG CCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGG GACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATC AAGTGTATCATATGCCAAGTCCGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGG CATTATGCCCAGTACATGACCTTACGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTC ATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACACCAATGGGCGTGGATAGCGGTTTGA CTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAA ATCAACGGGACTTTCCAAAATGTCGTAATAACCCCGCCCCGTTGACGCAAATGGGCGGTAG GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCGTTTAGTGAACCGTCAGATCACTAGAA GCTTTATTGCGGTAGTTTATCACAGTTAAATTGCTAACGCAGTCAGTGCTTCTGACACAACAG TCTCGAACTTAAGCTGCAGAAGTTGGTCGTGAGGCACTGGGCAGGTAAGTATCAAGGTTAC AAGACAGGTTTAAGGAGACCAATAGAAACTGGGCTTGTCGAGACAGAGAAGACTCTTGCGT TTCTGATAGGCACCTATTGGTCTTACTGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCAC TCCCAGTTCAATTACAGCTCTTAAGGCTAGAGTACTTAATACGACTCACTATAGGCTAGCATTT AGGTGACACTATAGAATACAAGCTACTTGTTCTTTTTGCACtcgaGgaattcGCCGCCACCATGAA CTCCTTCTCCACAAGCGCCTTCGGTCCAGTTGCCTTCTCCCTGGGCCTGCTCCTGGTGTTG CCTGCTGCCTTCCCTGCCCCAGTCTTCACACTCGAAGATTTCGTTGGGGACTGGCGACAGA CAGCCGGCTACAACCTGGACCAAGTCCTTGAACAGGGAGGTGTGTCCAGTTTGTTTCAGAA TCTCGGGGTGTCCGTAACTCCGATCCAAAGGATTGTCCTGAGCGGTGAAAATGGGCTGAAG ATCGACATCCATGTCATCATCCCGTATGAAGGTCTGAGCGGCGACCAAATGGGCCAGATCG AAAAAATTTTTAAGGTGGTGTACCCTGTGGATGATCATCACTTTAAGGTGATCCTGCACTATG GCACACTGGTAATCGACGGGGTTACGCCGAACATGATCGACTATTTCGGACGGCCGTATGA AGGCATCGCCGTGTTCGACGGCAAAAAGATCACTGTAACAGGGACCCTGTGGAACGGCAACAAAATTATCGACGAGCGCCTGATCAACCCCGACGGCTCCCTGCTGTTCCGAGTAACCATC AACGGAGTGACCGGCTGGCGGCTGTGCGAACGCATTCTGGCGGGTGGAGGCTTTGTGAA CCAACACCTGTGCGGCTCACACCTGGTGGAAGCTCTCTACCTAGTGTGCGGGGAACGAGG CTTCTTCTACACACCCAAGACCCGCCGGgAGGCaGAGGACCTGCAGGTGGGGCAGGTGGA GCTGGGCGGGggccCaggccCTGGTGCAGGCAGCCTGCAGCCCTTGGCCCTGGAGGGGTCC cTgcAGAAGCGTGGCATTGTGGAACAATGCTGTACCAGCATCTGCTCcctctaccagctggagaactac tgcaactagGAATTCcggtacCTCTAGAGTCGACCCGGGCGGCCGCTTCGAGCAGACATGATAAG ATACATTGATGAGTTTGGACAAACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGA AATTTGTGATGCTATTGCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACAACAACA ATTGCATTCATTTTATGTTTCAGGTTCAGGGGGAGATGTGGGAGGTTTTTTAAAGCAAGTAAA ACCTCTACAAATGTGGTAAAATCGATAAGGATCCGGGCTGGCGTAATAGCGAAGAGGCCCG CACCGATCGCCCTTCCCAACAGTTGCGCAGCCTGAATGGCGAATGGACGCGCCCTGTAGC GGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGC GCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTC CCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTC GACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGG TTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAA CAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTA TTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATTAACGCTTA CAATTTCCTGATGCGGTATTTTCTCCTTACGCATCTGTGCGGTATTTCACACCGCATATGGTG CACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGCCAGCCCCGACACCCGCCAACA CCCGCTGACGCGCCCTGACGGGCTTGTCTGCTCCCGGCATCCGCTTACAGACAAGCTGTG ACCGTCTCCGGGAGCTGCATGTGTCAGAGGTTTTCACCGTCATCACCGAAACGCGCGAGA CGAAAGGGCCTCGTGATACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGA CGTCAGGTGGCACTTTTCGGGGAAATGTGCGCGGAACCCCTATTTGTTTATTTTTCTAAATAC ATTCAAATATGTATCCGCTCATGAGACAATAACCCTGATAAATGCTTCAATAATATTGAAAAAGG AAGAGTATGAGTATTCAACATTTCCGTGTCGCCCTTATTCCCTTTTTTGCGGCATTTTGCCTT CCTGTTTTTGCTCACCCAGAAACGCTGGTGAAAGTAAAAGATGCTGAAGATCAGTTGGGTG CACGAGTGGGTTACATCGAACTGGATCTCAACAGCGGTAAGATCCTTGAGAGTTTTCGCCC CGAAGAACGTTTTCCAATGATGAGCACTTTTAAAGTTCTGCTATGTGGCGCGGTATTATCCCG TATTGACGCCGGGCAAGAGCAACTCGGTCGCCGCATACACTATTCTCAGAATGACTTGGTTG AGTACTCACCAGTCACAGAAAAGCATCTTACGGATGGCATGACAGTAAGAGAATTATGCAGT GCTGCCATAACCATGAGTGATAACACTGCGGCCAACTTACTTCTGACAACGATCGGAGGACC GAAGGAGCTAACCGCTTTTTTGCACAACATGGGGGATCATGTAACTCGCCTTGATCGTTGGG AACCGGAGCTGAATGAAGCCATACCAAACGACGAGCGTGACACCACGATGCCTGTAGCAAT GGCAACAACGTTGCGCAAACTATTAACTGGCGAACTACTTACTCTAGCTTCCCGGCAACAATTAATAGACTGGATGGAGGCGGATAAAGTTGCAGGACCACTTCTGCGCTCGGCCCTTCCGGC TGGCTGGTTTATTGCTGATAAATCTGGAGCCGGTGAGCGTGGGTCTCGCGGTATCATTGCA GCACTGGGGCCAGATGGTAAGCCCTCCCGTATCGTAGTTATCTACACGACGGGGAGTCAGG CAACTATGGATGAACGAAATAGACAGATCGCTGAGATAGGTGCCTCACTGATTAAGCATTGG TAACTGTCAGACCAAGTTTACTCATATATACTTTAGATTGATTTAAAACTTCATTTTTAATTTAAA AGGATCTAGGTGAAGATCCTTTTTGATAATCTCATGACCAAAATCCCTTAACGTGAGTTTTCG TTCCACTGAGCGTCAGACCCCGTAGAAAAGATCAAAGGATCTTCTTGAGATCCTTTTTTTCT GCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCG GATCAAGAGCTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAA TACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTA CATACCTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTT ACCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGG GGTTCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGC GTGAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAA GCGGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTAT CTTTATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTC AGGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGCCTTTTTACGGTTCCTGGCCTT TTGCTGGCCTTTTGCTCACATGGCTCGACAGATCT (SEQ ID NO: 97)

[0201] The recombinant antigen molecule of the present invention

[0202] In one aspect of the invention, the test kit for performing an assay according to the invention comprises a set of at least three recombinant antigen molecules in a single composition. The recombinant antigen molecules according to the invention are not particularly limited other than by their ability to be bound by antibodies, more specifically by antibodies against self-proteins (i.e., autoantibodies), preferably by islets autoantibodies, more preferably by autoantibodies against any of human insulin, proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, and human Zinc transporter 8.

[0203] In one embodiment, the set of at least three recombinant antigen molecules is selected from the group comprising recombinant human insulin human, recombinant human proinsulin, recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8. In another embodiment, the set of at least three recombinant antigen molecules is selected from the group consisting of human insulin human, recombinant human proinsulin, recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8. In a preferred embodiment, the recombinant antigen molecules are fused to a bioluminescent reporter protein.

[0204] In one embodiment, the bioluminescent reporter protein is a nanoluciferase reporter protein. In a preferred embodiment, the bioluminescent reporter protein is a Nanoluc luciferase (NLuc) reporterprotein. In a more preferred embodiment, the Nanoluc luciferase (NLuc) reporter protein comprises a nucleotide sequence at least 85%, preferably at least 90%, 95%, 98, 99% or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 10,.

[0205] It will be understood that the recombinant antigen molecules according to the invention are meant to optionally include a secretion signal peptide sequence. It is also understood that any reference to amino acid sequences referred to herein is meant to encompass not only the unmodified amino acid sequence but also typical posttranslational modifications of these amino acid sequences (e.g., glycosylation or deamidation of amino acids, the clipping of particular amino acids or other posttranslational modifications) occurring in cellular expression systems known in the art, including mammalian cells, such as Expi293F™ cells, or a sequence with one or more amino acid difference with the herein define amino acid sequence, such as 1, 2, 3, 4, 5, 6 or more amino acid difference.

[0206] The test kit for performing the assay according to the invention

[0207] In one aspect of the invention, the assay comprises a luciferase immune precipitation system (LIPS) assay or a solid phase capture LIPS (scLIPS) assay. Preferably, the assay is a luciferase immune precipitation system (LIPS) assay or a solid phase capture LIPS (scLIPS) assay The LIPS assay is well-known in the field of immune precipitation assays. The LIPS assay can be performed according to standard protocols described in the literature. LIPS is a type of liquid phase immunoassay that uses the emission of light from an enzymatic reaction to detect and quantify specific analytes in biological samples. A substrate of the reporter protein is added to the assay and the reporter protein catalyses a reaction that produces light. The amount of light produced can be detected and provides a quantitative measure of the level of the reporter protein in the sample; the level of the reporter protein in a sample may be used to determine the presence and / or level of an antibody molecule in a sample.

[0208] In one embodiment, the assay comprises or is a scLIPS assay is based on the LIPS assay and further comprises a protein selected from the group comprising protein A, protein L, anti IgA, anti IgM, and anti IgG. The protein is immobilized on a surface. As long as a surface can be coated with the protein that can bind an immune complex, any surface can be used. In one embodiment, the protein is immobilized on a surface selected from the group comprising a microplate, a particle, and resin. Preferably, the protein is immobilized on a surface selected from the group consisting of a microplate, a particle, and resin. When the surface is a particle, the particle is preferably a Sepharose bead, an agarose bead, or a paramagnetic particle. The size of the microplate is not particularly limited, and the type of surface can be different as long as it has the ability to be coated with a protein selected from the group comprising protein A, protein L, anti IgA, anti IgM, and anti IgG. For example, polystyrene plates with high binding capacity can be used, as well other typesof plates, such as plates with e.g., nylon membranes with a high binding capacity. In another embodiment, the protein is immobilized on the surface of a coated microfluidic device.

[0209] Advantageously, the LIPS assay and scLIPS assay offer high sensitivity, a wide dynamic range, and faster execution times compared to other immunoassay formats.

[0210] In one embodiment, the recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8 are diluted to a concentration of 2.0x106Lll / 25 pl (±200,000 LU) and combined to a final concentration of 6.0x106LU / 25 pl (±200,000 LU). In another embodiment, the human insulin and recombinant human proinsulin are diluted to a concentration of 10.0x106LU / 5 pl (±200,000 LU).

[0211] In a preferred embodiment, the recombinant human Glutamate decarboxylase 65, the recombinant human islet antigen 2, and the recombinant human Zinc transporter 8 molecules are mixed in a ratio of 1: 1: 1, preferably in a molar ratio of 1: 1: 1 in a single composition.

[0212] In one embodiment, the recombinant human insulin molecules, recombinant human proinsulin molecules, the recombinant human Glutamate decarboxylase 65 antigen molecules, the recombinant human islet antigen 2 antigen molecules, and the recombinant human Zinc transporter 8 antigen molecules are diluted in a buffer, including but not limited to TBST buffer or PBS. TBST buffer generally comprises 20 mM tris buffer, 150mM NaCI, and 0.5% Tween-20, at pH 7.4.

[0213] In one embodiment, the test kit further comprises at least three single compositions, wherein each single composition comprises a recombinant antigen molecule selected from the group comprising of recombinant human insulin, recombinant human proinsulin, recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8. In another embodiment, the test kit further comprises at least three single compositions, wherein each single composition comprises a recombinant antigen molecule selected from the group consisting of recombinant human insulin, recombinant human proinsulin, recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8. In a preferred embodiment, the recombinant antigen molecule is fused to a bioluminescent reporter protein. In a preferred embodiment, the bioluminescent reporter protein is a Nanoluc luciferase (NLuc) reporter protein. In a more preferred embodiment, the Nanoluc luciferase (NLuc) reporter protein comprises a nucleotide sequence at least 85%, preferably at least 90%, 95%, 98, 99% or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 10,.

[0214] In one embodiment, the recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8 are each diluted to a final concentration of about 4.0x106LU / 25pl (±200,000 LU) when present in three single compositions. In another embodiment, the recombinant human insulin and recombinant human proinsulin are each dilutedto a final concentration of about 10.0x106Lll / 5 l (±200,000 LU) when present in a single composition. In a preferred embodiment, the recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8 are each diluted to a final concentration of about 4.0x106LU / 25pl (±200,000 LU) and / or the recombinant human insulin and recombinant human proinsulin are each diluted to a final concentration of about 10.0x106LU / 5 pl (±200,000 LU) when present in at least three single compositions.

[0215] In an additional aspect of the invention, the test kit comprises the recombinant human insulin antigen molecule comprises a nucleotide sequence at least 85% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 1, and 11 to 46, at least 90%, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 1, and 11 to 46. In a preferred embodiment, the recombinant human insulin antigen molecule comprises a nucleotide sequence at least 85% identical to the nucleotide sequence encoded by SEQ ID NO: 1, at least 85%, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 1.

[0216] In an additional aspect of the invention, the test kit comprises the recombinant human proinsulin antigen molecule comprises a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 65 to 87 and 96. In an additional aspect of the invention, the test kit comprises the recombinant human islet antigen 2 antigen molecule comprises a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 2 to 4.

[0217] In an additional aspect of the invention, the test kit comprises the recombinant human Zinc transporter 8 antigen molecule comprises a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 5.

[0218] In an additional aspect of the invention, the test kit comprises the recombinant human Glutamate decarboxylase 65 antigen molecule comprises a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 6.

[0219] In one aspect of the invention, the recombinant antigen molecules of the kit are suitable for the detection of human autoantibody molecules, preferably islets autoantibodies in a sample obtained from a human, preferably selected form: a blood sample, a serum sample and a plasma.

[0220] The kit of the present invention is suitable for the identification of at least one autoantibody molecule with reactivity to a self-antigen, such as an islet autoantibody molecule, in a patient developing type 1 diabetes. Preferably, the autoantibody molecule is at least one autoantibody molecule with specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, or human Zinc transporter 8. More preferably, the antibody is at least one islet autoantibody selected from the group comprising islet autoantibodies IAA (Insulin Auto Antibodies), GADA (Glutamic Acid Decarboxylase Antibodies), IA-2A (Islet Antigen 2 Antibodies),and ZnT8A (Zinc Transporter 8 Antibodies). Most preferably, the autoantibody is at least one islet autoantibody selected from the group consisting of islet autoantibodies IAA (Insulin Auto Antibodies), GADA (Glutamic Acid Decarboxylase Antibodies), IA-2A (Islet Antigen 2 Antibodies), and ZnT8A (Zinc Transporter 8 Antibodies).

[0221] Hence, the present invention provides a kit for the in vitro diagnosis of type 1 diabetes.

[0222] The present invention allows for a rapid, sensible and efficient identification of the presence of autoantibodies, such as islet autoantibodies in a sample, which may be indicative of the presence of an autoimmune disorder, such a type 1 diabetes.

[0223] In certain embodiments, the kit may also comprise a sterile container such as boxes, ampules, bottles, vials, tubes, bags, pouches, blister-packs, dishes or other suitable container forms known in the art. Such containers can be made of plastic, glass or other materials suitable for holding cells. The kit may also comprise suitable cell culture medium, shipping medium, buffers and any other reactive that may become necessary for detecting autoantibody molecules.

[0224] In one aspect of the invention, the recombinant antigen molecules, or the test kit according to the present invention are meant to be used in a test at entry into a screening program. The assay comprising the recombinant antigen molecules or the test kit according to the invention can be followed by repeat confirmatory tests in which samples positive in the assay of the invention are re-analyzed to confirm the presence and type of an antibody in the sample. In one embodiment, the tests to confirm the presence and type of an antibody present in the sample comprise a recombinant antigen molecule selected from the group consisting of recombinant human insulin, recombinant human proinsulin, recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8 according to the invention. In one embodiment, the tests to confirm the presence and type of an antibody present in the sample comprise an immunoassay, such as a LIPS assay.

[0225] The nucleic acid, vector, and cell of the present invention

[0226] In a further aspect, the invention provides a nucleic acid encoding the recombinant antigen molecule of the test kit. In a preferred embodiment, each of the recombinant antigen molecule is encoded by a separate nucleic acid. In another preferred embodiment, each of the recombinant antigen molecule is encoded by separate vector. In one embodiment, the vector is a lentiviral vector. In another embodiment, the vector is a pCMV plasmid. In a further embodiment, the vector is a pCMVTnT plasmid.

[0227] The invention also provides a vector encoding for:

[0228] a. a recombinant human insulin antigen molecule comprising a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ I D NOs: 1, 11 to 46, and 47 to 60,; orb. a recombinant human proinsulin antigen molecule comprising a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 62, 63, 65 to 87, and 88 to 96,; or

[0229] c. a recombinant human islet antigen 2 antigen molecule comprising a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 2 to 4; or

[0230] d. a recombinant human Zinc transporter 8 antigen molecule comprising a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 5; or

[0231] e. a recombinant human Glutamate decarboxylase 65 antigen molecule comprising a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 6.

[0232] In a preferred embodiment, the recombinant human insulin antigen molecule comprises a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 1.

[0233] In one embodiment, the invention provides a cell comprising the set of nucleic acids or a nucleic acid, or a vector and expressing the recombinant antigen molecules. In another preferred embodiment, the cell is a stably expressing transfectant cell line. In one preferred embodiment, the cell is a mammalian cell, such as an Expi293F™ cell. In one embodiment, the cell expresses a nucleic acid or a vector encoding for one recombinant antigen molecule selected from the group comprising recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin, and recombinant human proinsulin. In a more preferred embodiment, the cell expresses a nucleic acid or a vector encoding for one recombinant antigen molecule selected from the group consisting of recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin, and recombinant human proinsulin. In another embodiment, the transfectant cell line is an Expi-293F NLuc GAD65 aa96-585 cell line, an Expi-293F Dual NLuc I A-2ic cell line, an Expi-293F Dual NLuc ZnT8 dimer cells line, or an Expi-293F Insulin B NLuc cell line. Advantageously, the stably expressing transfectant cell line facilitates cheaper and larger scale production of the recombinant antigen molecules according to the invention.

[0234] The method of the present invention

[0235] In a further aspect, the invention provides a method for producing the recombinant antigen molecule of the test kit, the method comprising expressing the set of nucleic acids or the nucleic acid or the vector in a cell, and harvesting the recombinant antigen molecule.The recombinant antigen molecules according to the invention can be obtained by methods known in the art. Such methods include methods for the production of recombinant antigen molecules, e.g., of recombinant proteins.

[0236] In another aspect, the invention provides a method for determining the presence and / or level of an antibody molecule in a sample.

[0237] According to the invention, the present invention provides an in vitro method for determining the presence and / or level of an antibody molecule in a sample. In one aspect, the sample is a blood sample, a serum sample, or a plasma sample obtained from a human subject.

[0238] In some embodiments, the sample is provided in a form requiring reconstitution, preferably as a dry sample that is reconstituted prior to the step of contacting the sample with the at least three antigens. In certain embodiments, the sample is a dried blood spot (DBS), preferably, collected on a substrate, e.g. a filter paper and reconstituted prior to contacting the sample with the at least three antigens. In some embodiments, the sample is lyophilized serum or plasma to be reconstituted before the assay.

[0239] In some embodiments, the sample is reconstituted, prior the phase of contacting with the at least three antigens, in a liquid, preferably in a salt buffer. In certain embodiments, the liquid comprises TBST buffer, optionally supplemented with fetal bovine serum (FBS). In some embodiments, the buffer comprises a high-salt concentration to reduce nonspecific binding.

[0240] The in vitro method comprises the steps of:

[0241] a) contacting the sample with the set of at least three recombinant antigen molecules, wherein a complex between the recombinant antigen molecule and the autoantibody molecule (referred to as “antigen molecule-autoantibody complex”) is formed;

[0242] b) contacting the recombinant antigen molecule-autoantibody complex with a protein; c) removing the unbound recombinant antigen molecules; and

[0243] d) contacting the bioluminescence reporter protein covalently linked to the recombinant antigen molecule with the substrate;

[0244] e) determining the presence, absence, and / or amount of one or more autoantibody molecules in the sample that binds to one or more recombinant antigen molecules.

[0245] In one embodiment of step a), the sample is contacted with a single composition comprising recombinant human Glutamate decarboxylase 65 antigen molecule, recombinant human islet antigen 2 antigen molecule, recombinant human Zinc transporter 8 antigen molecule, recombinant human insulin antigen molecule, and recombinant human proinsulin antigen molecule. In a preferred embodiment of step a), the sample is contacted with a single composition comprising recombinant human Glutamate decarboxylase 65 antigen molecule, recombinant human islet antigen 2 antigen molecule, and recombinant human Zinc transporter 8 antigen molecule. The skilled person is able to determine the conditions and the length of the contactingperiod which are sufficient for the formation of a complex between the recombinant antigen molecule and the autoantibody molecule (referred to as “antigen molecule-autoantibody complex”). Suitable buffers, salts, solutions, or in culture medium are comprised in the single composition comprising the at least three recombinant antigen molecules.

[0246] In one embodiment of step b), the recombinant antigen molecule-autoantibody complex is contacted with a protein selected from the group comprising protein A, protein L, anti IgA, anti IgM, and anti IgG. In another embodiment, the recombinant antigen molecule-autoantibody complex is contacted with a protein selected from the group consisting of protein A, protein L, anti IgA, anti IgM, and anti IgG. In a preferred embodiment, the protein is immobilized on a surface selected from the group comprising a microplate, a particle, and resin. In another preferred embodiment, the protein is immobilized on a surface selected from the group consisting of a microplate, a particle, and resin. Preferably, the particle is a Sepharose bead, an agarose bead, or a paramagnetic particle.

[0247] In one embodiment of step c), the removal of unbound recombinant antigen molecules may be achieved by washing to eliminate any unbound recombinant antigen molecules. Preferably, this step is performed to remove unbound recombinant antigen molecules and unbound autoantibody molecules. Most preferably, the recombinant antigen molecule-autoantibody complex is retained and subject to the subsequent step.

[0248] In one embodiment of step d), the bioluminescence reporter protein covalently linked to the recombinant antigen molecule is contacted with a substrate for the bioluminescence reporter protein. Suitable substrates for the bioluminescence reporter protein according to the invention are well-known in the field of immunoassays. The substrate might comprise further component and buffers required for the enzymatic activity of the bioluminescent reporter protein.

[0249] In one embodiment of step e), the presence, absence, or amount of one or more autoantibody molecules in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter molecule.

[0250] In one embodiment, the sample is further contacted with at least three recombinant antigen molecules in at least three single compositions. Preferably, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions when the sample is found positive for at least one autoantibody molecules as determined in step e).

[0251] Accordingly, and subsequently to the method steps a) to e) as described above, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions, wherein the method comprises the steps of:

[0252] f) contacting the sample with the set of at least three recombinant antigen molecules in at least three single compositions, wherein a recombinant antigen molecule-autoantibody complex is formed;g) contacting the recombinant antigen molecule-autoantibody complex with a protein; h) removing the unbound recombinant antigen molecules;

[0253] i) contacting the bioluminescence reporter protein covalently linked to the recombinant antigen molecule with the substrate; and

[0254] j) determining the presence, absence, and / or amount of one or more autoantibody molecules in the sample that binds to any of the at least three recombinant antigen molecule,

[0255] wherein the presence, absence, or amount of one or more autoantibody molecules in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter molecule. In one embodiment, the at least one autoantibody molecule is against a self-antigen. Preferably, the at least one autoantibody molecule is an islet autoantibody molecule, optionally wherein the islet autoantibody molecule has specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, or human Zinc transporter 8. More preferably, the islet autoantibody molecule is selected from the group comprising or consisting of islet autoantibodies IAA (Insulin Auto Antibodies), GADA (Glutamic Acid Decarboxylase Antibodies), IA-2A (Islet Antigen 2 Antibodies), and ZnT8A (Zinc Transporter 8 Antibodies).

[0256] In one aspect, the assay according to the invention comprises a luciferase immune precipitation system (LIPS) assay or a modified LIPS assay, referred to as solid-phase capture LIPS (scLIPS) assay. The LIPS assay is well-known in the field of immune precipitation assays. The LIPS assay can be performed according to standard protocols described in the literature. LIPS is a type of liquid phase immunoassay that uses the emission of light from an enzymatic reaction to detect and quantify specific analytes in biological samples. A substrate of the reporter protein is added to the assay and the reporter protein catalyses a reaction that produces light. The amount of light produced can be detected and provides a quantitative measure of the level of the reporter protein in the sample; the level of the reporter protein in a sample may be used to determine the presence and / or level of an antibody molecule in a sample. In one embodiment, the assay comprises a scLIPS assay is based on the LIPS assay and further comprises a protein selected from the group comprising protein A, protein L, anti IgA, anti IgM, and anti IgG.

[0257] Advantageously, the LIPS assay and scLIPS assay offer high sensitivity, a wide dynamic range, and faster execution times compared to other immunoassay formats.

[0258] In another aspect, the in vitro method comprises or is a luciferase immune precipitation system (LIPS) assay. In a preferred embodiment, the method comprises the steps of:

[0259] a) contacting the sample with the set of at least three recombinant antigen molecules; b) contacting the recombinant antigen molecule-autoantibody complex with a protein immobilized on a Sepharose bead,, an agarose bead, a paramagnetic particle, or resin; c) removing the unbound recombinant antigen molecules;d) contacting the bioluminescence reporter protein covalently linked to the recombinant antigen molecule with the substrate; and

[0260] e) determining the presence, absence, and / or amount of one or more autoantibody molecules in the sample that binds to one or more recombinant antigen molecules, wherein the presence, absence, or amount of one or more autoantibody molecules in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter molecule. According to the invention, the LIPS assay can be performed according to standard protocols described in the literature.

[0261] In one embodiment of step a), a single-use antigen aliquots of the recombinant antigens are thawed at room temperature and diluted in Tris-buffered saline pH 7.4 with 0.5% Tween 20 (TBST-0.5%) to achieve a concentration for each antigen of 2.0x106Lll / 25 pl (±200,000 LU). Antigens are then combined to a yield a cumulative luciferase activity of 6.0x106LU / 25 pl (±200,000 LU) and filtered through a PVDF 0.45pM Millex-SV syringe filter. The total volume of diluted antigen to be prepared is determined by the number of samples to be tested. In one embodiment, 1 pl of the sample, preferably serum, is pipetted into a well of a 96-deep well polystyrene.

[0262] In one embodiment of step b), the recombinant antigen molecule-autoantibody complex is contacted with 5pL of blocked rProtein A Sepharose 4Fast Flow resuspended in 50pL, for 1h at 4 °C with shaking. In one embodiment, a PAS 50% stock slurry in EtOH 20 is prepared by blocking with glycine. Before use in the LIPS assay, 5 pl / well of PAS is washed four times in TBST and diluted in TBST to a final volume of 50 pl / well and then added to each well of the incubation plates. Plates are then incubated for 1 hr with orbital shaking (-700 rpm) at 4°C.

[0263] In one embodiment of step c), the unbound recombinant antigen is removed by serial washes. The washes can comprise at least 1, 2, 3, 4, 5 serial washes, preferably, 5 serial washes. The washes can be performed with a suitable buffer, such as for example TBST. In one example, each wash consists in the addition of 750 pl / well of TBST, followed by the plate centrifugation (e.g., 500xg for 3 min at 4°C) and buffer removal e.g., by aspiration using a micro-plate plate washer. In one embodiment of step d) and step e), the resin pellets are transferred to an OptiPlate™ and the bioluminescence activity, e.g., the luciferase activity, is measured upon addition of 20pL to 40pL of Nano-Gio® substrate and 20 of TBST to each well followed by immediate readout in a luminometer. In one embodiment, 0.5X Nano-Gio® Luciferase Assay Reagent is prepared in advance by bringing at room temperature NanoGio® Luciferase Assay Buffer (stored frozen at -20° C). To measure the luciferase activity, the required reagents are prepared by combining 0.4 ml of Nano-Gio® Luciferase Assay Substrate (which is a liquid stored at -20°C), 20 ml of Nano-Glo® Luciferase Assay Buffer, and 20 ml of TBST previously equilibrated to RT per well. Using a multichannel dispenser, 40 pl of 0.5X Nano-Gio® substrate is added to each well of the OptiPlates™ followed by immediate readout in Centro X960 luminometer using a protocolconsisting of shaking for 1 seconds followed by light acquisition for 2 seconds / well. Acquired LU are then converted to arbitrary units (AU) using a standard curve constructed from doubling dilutions in normal human serum of a multiple GADA, IA-2A and ZnT8A Ab positive serum. AU are calculated based on the measured LU, using a mixed Iog10 (for light units) and Iog2 (for standard concentrations) logarithmic curve fitting algorithm in Excel (Microsoft Corporation).

[0264] In one embodiment, the presence of antibody molecule with specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, and / or human Zinc transporter 8 in the sample is indicative of a risk to develop type 1 diabetes.

[0265] Advantageously, the LIPS assay requires an extremely limited amount of serum or plasma for each test. In one embodiment of the present invention, only 1 microliter of serum per replicate is required for the LIPS assay according to the invention, whereas 2-5 microliters are required in the RBA and 25 microliters in the RSR bridge-ELISA. For example, for an IAA LIPS assay, performed with and without cold insulin competition in 4 replicate wells, the total serum consumption is 4 microliters.

[0266] A further advantage of the present invention relates to the relatively short assay duration. For example, the incubation time for the GADA, IA-2A, ZnT8A LIPS assay is 2 hours, compared to an overnight incubation for RBA or RSR bridge-ELISA. Similarly, the incubation time for the IAA LIPS assay is of 18 hours compared to 72 hours for the RBA.

[0267] In one embodiment, the sample is further contacted with at least three recombinant antigen molecules in at least three single compositions. Preferably, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions when the sample is found positive for at least one autoantibody molecules as determined in step e).

[0268] Accordingly, and subsequently to the method steps a) to e) as described above, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions, wherein the method comprises the steps of:

[0269] f) contacting the sample with the set of at least three recombinant antigen molecules in at least three single compositions, wherein a recombinant antigen molecule-autoantibody complex is formed;

[0270] g) contacting the recombinant antigen molecule-autoantibody complex with a protein; h) removing the unbound recombinant antigen molecules;

[0271] i) contacting the bioluminescence reporter protein covalently linked to the recombinant antigen molecule with the substrate; and

[0272] j) determining the presence, absence, and / or amount of one or more autoantibody molecules in the sample that binds to any of the at least three recombinant antigen molecule,wherein the presence, absence, or amount of one or more autoantibody molecules in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter molecule. In another aspect of the invention, the presence, absence, or amount of one or more antibody molecules in the sample is determined with a solid phase capture LIPS (scLIPS) assay. Accordingly, in one embodiment, the in vitro method comprises a solid phase capture luciferase immune precipitation system (scLIPS) assay. The scLIPS assay can be performed by performing the LIPS assay, wherein the recombinant antigen molecule-antibody complex is bound by a protein selected from the group comprising protein A, protein L(e.g., this recognizes the light chain of an antibody and can thus recognize several antibody types), anti IgA, anti IgM, and anti IgG prior to measuring the bioluminescence activity.

[0273] In a preferred embodiment, the method comprises the steps of:

[0274] a) contacting the sample with the set of at least three recombinant antigen molecules, wherein a recombinant antigen molecule-autoantibody complex is formed;

[0275] b) contacting the recombinant antigen molecule-autoantibody complex with a protein immobilized on a surface;

[0276] c) removing the unbound recombinant antigen molecules;

[0277] d) contacting the bioluminescence reporter protein covalently linked to the recombinant antigen molecule with the substrate; and

[0278] e) determining the presence, absence, and / or amount of one or more autoantibody molecules in the sample that binds to one or more recombinant antigen molecules, wherein the presence, absence, or amount of one or more autoantibody molecules in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter molecule. In one embodiment of step a), 1 l of the sample, preferably the serum, is pipetted to a polypropylene 96-well plate. 25pl of the composition comprising the at least three recombinant antigen molecules are added, briefly mixed on a plate mixer, and incubated for 2 hr at room temperature protected from light.

[0279] In one embodiment of step b), the protein is selected from the group comprising or consisting of protein A, protein L, anti IgA, anti IgM, and anti IgG. In one embodiment, the protein binds to any of the component of the complex. In a preferred embodiment, the protein binds to the constant portion (e.g., the Fc portion) of the autoantibody molecule. In a preferred embodiment, the recombinant antigen molecule-antibody complex is bound by Protein A and captured on an immunoglobulin G-binding protein-A-coated plate prior to measuring the bioluminescence activity, e.g., preferably the Nanoluc luciferase (NLuc) activity. In one embodiment, the reaction mixture of step a) is transferred to 96-well plates pre-coated with 30 mg / well of recombinant Protein A in carbonate buffer pH9 for 4 hours at room temperature and then blocked with freshly prepared 1,5% Fish Gelatin in PBS-HS 1X plus 0,1% Tween-20 pH9 for 18 hours at 4°C. Alternatively,commercial plates can be used coated with recombinant protein A / G and blocked with BSA. Capture plates are incubated for 1 hr with orbital shaking (-700 rpm) at 4°C.

[0280] In one embodiment of step c), the unbound recombinant antigen molecules are washed e.g., with TBT using the BioTek Elx405 performing 10 washes of 300 ml each.

[0281] In one embodiment of step d) and step e), 0.5X Nano-Gio® Luciferase Assay Reagent is prepared in advance by bringing at room temperature NanoGio® Luciferase Assay Buffer (stored frozen at -20° C). To measure the luciferase activity, the required reagents are prepared by combining 0.4 ml of Nano-Gio® Luciferase Assay Substrate (which is a liquid stored at -20°C), 20 ml of Nano-Glo® Luciferase Assay Buffer, and 20 ml of TBST previously equilibrated to RT per well. Using a multichannel dispenser, 40 pl of 0.5X Nano-Gio® substrate is added to each well of the OptiPlates™ followed by immediate readout in Centro X960 luminometer using a protocol consisting of shaking for 1 seconds followed by light acquisition for 2 seconds / well. Acquired LU are then converted to arbitrary units (AU) using a standard curve constructed from doubling dilutions in normal human serum of a multiple GADA, IA-2A and ZnT8A Ab positive serum. AU are calculated based on the measured LU, using a mixed Iog10 (for light units) and Iog2 (for standard concentrations) logarithmic curve fitting algorithm in Excel (Microsoft Corporation).

[0282] In one embodiment, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions. Preferably, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions when the sample is found positive for at least one autoantibody molecule as determined in step e).

[0283] Accordingly, and subsequently to the method steps a) to e) as described above, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions, wherein the method comprises the steps of:

[0284] f) contacting the sample with the set of at least three recombinant antigen molecules in at least three single compositions, wherein a recombinant antigen molecule-autoantibody complex is formed;

[0285] g) contacting the recombinant antigen molecule-autoantibody complex with a protein; h) removing the unbound recombinant antigen molecules;

[0286] i) contacting the bioluminescence reporter protein covalently linked to the recombinant antigen molecule with the substrate; and

[0287] j) determining the presence, absence, and / or amount of one or more autoantibody molecules in the sample that binds to any of the at least three recombinant antigen molecule,

[0288] wherein the presence, absence, or amount of one or more autoantibody molecules in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter molecule.In one embodiment, the sample is contacted with the set of at least three recombinant antigen molecules in a single composition. In a preferred embodiment, the single composition comprises recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, and recombinant human Zinc transporter 8. In a more preferred embodiment, the single composition does not comprise recombinant human insulin and / or recombinant human proinsulin. In a most preferred embodiment, the sample is further contacted with recombinant human insulin and / or recombinant human proinsulin in a single composition.

[0289] In another embodiment, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions. Preferably, the sample is contacted with at least three recombinant antigen molecules in at least three single compositions when the sample is found positive for at least one autoantibody molecules when contacted with the set of at least three recombinant antigen molecules in a single composition and / or when contacted with recombinant human insulin in a single composition and / or recombinant human proinsulin in a single composition.

[0290] In one embodiment, the presence of antibody molecule with specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, and / or human Zinc transporter 8 in the sample is indicative of a risk to develop type 1 diabetes.

[0291] Advantageously, the scLIPS assay requires an extremely limited amount of serum or plasma for each test. In one embodiment of the present invention, only 1 microliter of serum per replicate is required for the scLIPS assay according to the invention, whereas 2-5 microliters are required in the RBA and 25 microliters in the RSR bridge-ELISA. For example, for an IAA scLIPS assay, performed with and without cold insulin competition in 4 replicate wells, the total serum consumption is 4 microliters.

[0292] A further advantage of the present invention relates to the relatively short assay duration. For example, the incubation time for the GADA, IA-2A, ZnT8A scLIPS assay is 2 hours, compared to an overnight incubation for RBA or RSR bridge-ELISA. Similarly, the incubation time for the IAA scLIPS assay is of 18 hours compared to 72 hours for the RBA.

[0293] In another aspect, the autoantibody molecule is specific to an antigen expressed by a human beta cell. In one embodiment, the autoantibody molecule is at least one autoantibody molecule against a self-antigen. In another embodiment, the autoantibody molecule is at least one islet autoantibody molecule. In further embodiment, the islet autoantibody is selected from the group comprising the islet autoantibodies IAA (Insulin Auto Antibodies), GADA (Glutamic Acid Decarboxylase Antibodies), IA-2A (Islet Antigen 2 Antibodies), and ZnT8A (Zinc Transporter 8 Antibodies). In a preferred embodiment, the islet autoantibody is at least one autoantibody molecule with specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, or human Zinc transporter 8.In one aspect, the method comprises screening a human subject for the development of type 1 diabetes based on the presence, absence, or amount of one or more antibody molecules in the sample, wherein the presence, absence, or amount of one or more antibody molecules in the sample is determined by comparing the amount of detected antibody molecule in the sample with a standard curve for known amounts of said antibody molecules. In one embodiment, the presence of one or more antibody molecules in the sample is indicative of a risk to develop type 1 diabetes.

[0294] In a further aspect, contacting of the autoantibodies present in the sample is performed by seeding 1-pL replicates of each sample into 96-deep-well plates and adding from 6pL to 25pL of a composition comprising the recombinant antigen molecules according to the invention. In one embodiment, the plates are incubated for 2h at room temperature after mixing and centrifugating. In one embodiment, the recombinant antigen molecule-autoantibody complex are captured by incubation with 5pL of blocked rProtein A Sepharose 4Fast Flow resuspended in 50pL, for 1h at 4 °C with shaking. In one embodiment, plates are washed 5 times by sequential dispensing of 750pL of TBST per well, centrifugation at 500g for 3 minutes at 4 °C, and removal of supernatant using a micro-plate plate washer. In one embodiment, the resin pellets are transferred to an OptiPlate™ and the luciferase activity is measured upon addition of 20pL to 40pL of Nano-Gio® substrate and 20 of TBST to each well followed by immediate readout in a luminometer. In one embodiment, the readout is performed using a protocol consisting of shaking for 1 second followed by light acquisition for 2 seconds / well in a luminometer.

[0295] In one embodiment, acquired LU are converted to arbitrary units (AU) using a standard curve constructed from doubling dilutions in healthy human sample of an IAA-, GADA-, IA-2A- orZnT8A-positive sample. In one embodiment, AU are calculated based on the measured LU, using a mixed Iog10 (for light units) and Iog2 (for standard concentrations) a logarithmic curve fitting algorithm. In one embodiment, the presence of at least one autoantibody molecule with specificity to human insulin, human proinsulin, human Glutamate decarboxylase 65, human islet antigen 2, and / or human Zinc transporter 8 in the sample is indicative of a risk to develop type 1 diabetes.

[0296] The LIPS and scLIPS assays according to the invention require an extremely limited amount of serum or plasma for each test. In one embodiment of the present invention, only 1 microliter of serum per replicate is required for the LIPS assay according to the invention, whereas 2-5 microliters are required in the RBA and 25 microliters in the RSR bridge-ELISA. For example, for an IAA LIPS assay, performed with and without cold insulin competition in 4 replicate wells, the total serum consumption is 4 microliters.

[0297] A further advantage of the present invention relates to the relatively short assay duration. For example, the incubation time for the GADA, IA-2A, ZnT8A LIPS assay is 2 hours, compared to anovernight incubation for RBA or RSR bridge-ELISA. Similarly, the incubation time for the IAA LIPS assay is of 18 hours compared to 72 hours for the RBA.

[0298] The presence and / or level of an autoantibody molecule in a sample may be assessed by comparing the level of autoantibody molecules in a sample from a subject at risk of developing T1 D, to the level of autoantibody molecules in a control sample, i.e., a sample from a subject that is not at risk of developing T1 D, wherein an increase level of autoantibody molecules indicates an increased probability of developing T1 D. Preferably, the level is statistically significant as assessed by appropriate statistical tests which are known in the art.

[0299] The present invention also provides a method of treating an autoimmune disorder, preferably an autoimmune disorder associated with the presence of islets autoantibodies, more preferably type 1 diabetes, comprising:

[0300] administering a treatment to effectively treat the autoimmune disorder in a subject who has been diagnosed as having an autoimmune disorder based on the presence of islets autoantibodies present in a sample obtained from the subject, wherein the islets autoantibodies bind to the recombinant antigen molecules of to the present invention.

[0301] MATERIALSAND METHODS

[0302] Cloning of antigens in plasmid vectors

[0303] The coding sequences of human T1D autoantigens were obtained starting from UniProt (www.uniprot.org) reference sequences. Modified coding sequences were amplified by RT-PCR from human islets cDNA and then mutagenized for GAD65 (UniProt entry Q05329), IA-2 (Uniprot entry Q16849) and ZnT8 (UniProt entry Q8IWU4). The antigen coding sequence of GADA65 was truncated to remove the first 95 amino acids that contains an epitope recognized by autoantibody reactivities poorly predictive of T1D development in pre-diabetes subjects. The IA-2 intracellular (IA-2ic) coding sequence was truncated to remove the first 604 amino acids, corresponding to extracellular and transmembrane domains and 1) do not contain main IA-2A epitopes and have 2) previously shown to be associated with higher unspecific background in immunoassays. The ZnT8 sequence was truncated to remove the first 267 amino acids which do not contain major epitopes of ZnT8A and to remove the transmembrane domains previously shown to be associated with higher unspecific background in immunoassays. Two distinct polymorphic variants were cloned containing a different polymorphic residue in amino acid 365 and encoding for a tryptophan or arginine residue, respectively. The two polymorphic ZnT8 COOH domains were then cloned in frame to create a dimeric ZnT8 recombinant antigen. The full-length human pre-proinsulin cDNA sequence was obtained as a synthetic chimeric DNA sequence in which a nanoluciferase reporter was inserted at the end of the insulin B chain before the C peptide and in which the proconvertase 2 recognition sequences in the C peptide were mutagenized to transform them into furin convertase recognition sequences. Several alternative variants of NanoLuc tagged proinsulin andinsulin recombinant antigens were produced. In particular, the impact on antigen expression and autoantibody binding of the luciferase reporter tag placement relative to the autoantigen coding sequence was initially investigated. Subsequently, the impact on recombinant protein expression and autoantibody binding of introduction of several deletion or point mutations on a selection of these human recombinant insulin antigens and proinsulin antigens used as backbones was also investigated (see Tables 2 to 4, and 6 to 10 containing the sequences of such recombinant antigens).

[0304] Site-directed mutagenesis of coding sequences to change specific amino acid residues or to introduce restriction sites at desired location in the coding sequence or to produce truncated forms was performed using the GeneArt® Site-Directed Mutagenesis PLUS Kit protocol (Thermo Fisher Scientific, Carlsbad, CA, USA).

[0305] All modified coding sequences were sub-cloned into modified pCMVTNT™ plasmid (Promega, Madison Wl, USA) expression vectors in which a NanoLuc™ (NLuc) luciferase reporter (Promega) was previously inserted, except for the insulin antigen construct. In the case of the truncated GAD65 antigen, the NanoLuc reporter was placed in frame upstream of the antigen coding sequence and joined to it by a glycine spacer. In the case of the I A-2ic cDNA sequence, this was cloned in frame both upstream and downstream of the NanoLuc reporter to generate a “dual” construct in which a dimeric intracellular IA-2 was joined by the interspersed reporter and separated by it by linker and spacer sequences. For NanoLuc tagged ZnT8 antigen a similar approach was used to generate a recombinant antigen in which two copies of the fused polymorphic domains were joined in frame both upstream and downstream of the luciferase reporter, respectively. This construct yielded a “dual” ZnT8 dimer luciferase tagged antigen. Expression and preparation of recombinant luciferase tagged antigens

[0306] Each recombinant Nanoluc® luciferase tagged antigens was initially expressed and validated using the Expi293™ Expression System (Thermo Fisher Scientific Life Technologies). Expression of each cDNA construct subcloned in the pCMVTnT™ plasmid (Promega, USA) was driven by the plasmid CMV promoter after transient transfection of Expi293F™ cells using Expifectamine (see Tables 5 and 10 which display the sequences of mutated insulin and proinsulin subcloned into the pCMVTnT™ plasmid). Briefly, for a typical small-scale transfection, Expi293F™ cells were grown until an early logarithmic growth phase was reached and counted in a Burker counting chamber. Then, 4.5 x 106Expi293F™cells were resuspended in 2.125 ml of Expi293F Expression Medium and seeded into the well of a 6-well cell culture plate. A transfection mix was then prepared in three steps: 1) the combination of 4 pg of plasmid in 125 pl of Opti-MEM I followed by incubation at room temperature for 5 minutes; 2) the combination of 6.8 ml of ExpiFectamine 293TM Reagent in 118 ml of Opti-MEM I Reduced Serum Medium followed by incubation at room temperature for 5 minutes; 3) the combination of the previous two mixes followed by incubation at roomtemperature for 20 minutes to allow the formation of plasmid and transfection agents complexes. Seeded Expi293F™ cells in the 6-well plate were then added with the transfection mix and incubated for up to 24 hours on an orbital shaker (set at 125 rpm) placed inside a CO2 cell culture incubator. Approximately 20 hours post-transfection, the transfected Expi293F™ cells were added first with 12.5 ml of Expifectamine 293 Transfection Enhancer 1 and then with 125 ml of Expifectamine 293 Transfection Enhancer 2 reagents followed by incubation of the 6-well plate for 24 hours on an orbital shaker (set at 125 rpm) placed inside a CO2 cell culture incubator. Approximately 48 hours post- transfection, transfected cells were transferred into a 2ml Eppendorf® LoBind microcentrifuge tube and centrifuged for 5 minutes at 100xg at room temperature. Depending on the antigen being secreted or not the antigen harvest follows a different procedure.

[0307] For cytoplasmic antigens like GAD65, IA-2, and ZnT8 the supernatant is removed, and the cell pellet washed with 2 ml of phosphate buffered saline (PBS) pH 7.4 at room temperature, followed by centrifugation for 5 minutes at 100xg at room temperature. The PBS is then discarded, and the pellet is resuspended by pipetting in 800 ml of Passive Lysis Buffer 1X (PLB) (Promega) to lyse cells and release proteins. The cell lysate is then transferred into a 2 ml low protein binding 2 ml tube (LoBind™ Eppendorf) and kept on ice for 10 minutes, this step include vortex thoroughly every 2-3 minutes. Finally, the tube is centrifuged at 13,000 rpm in a refrigerated minicentrifuge at 4°C for 10 minutes. The supernatant is collected taking care to avoid dislodging and transferring insoluble debris and dispensed into PCR strip tubes as single use 5 ml aliquots or alternatively into low protein binding Eppendorf tubes for larger aliquots. All processed aliquots are then stored immediately at -80°C. For secreted antigens, the expressed antigens are present in the transfected cells culture supernatant. The supernatant is collected and transferred to a new Eppendorf tube and centrifuged at 13,000 rpm in a refrigerated minicentrifuge at 4°C for 10 minutes to pellet insoluble debris. The supernatant is then dispensed into PCR strip tubes as single use 5 ml aliquots, or alternatively into low protein binding Eppendorf tubes for larger aliquots.

[0308] Estimate of Luciferase activity

[0309] After the harvest of expressed antigen and prior to -80°C storage, an estimate of the antigen content is obtained by determining the recovered luciferase activity in the processed cell lysate or the cleared supernatant, for cytoplasmic antigens or secreted antigens respectively. The procedure requires the thaw of NanoGio® Luciferase Assay Buffer (stored frozen at -20° C) that is then brought at room temperature. For each individual measurement of luciferase activity, the reconstituted Nano-Gio® Luciferase Assay Reagent is prepared by combining 0.8 ml of Nano-Glo® Luciferase Assay Substrate (which is a liquid stored at -20°C) with 40 ml of Nano-Gio® Luciferase Assay Buffer (previously equilibrated to RT). Harvested antigen (2 ml) is serially diluted1:250, 1:2500, 1:25000. Of each dilution, 25 ml are taken and mixed with 40 ml of Nano-Gio® Luciferase Assay Reagent and read into a luminometer (Centro X960, Berthold Germany) for 2 seconds. The luciferase activity is measured as light units (LU), i.e., single photon counting, and the actual emission due to the antigen-luciferase present in the preparation is converted to light units / ml by multiplying for the appropriate corrected dilution factor measurements within the linear range of the luminometer readout.

[0310] Generation of stable cell lines expressing luciferase tagged T1D autoantigens Subcloning of antigen-luciferase coding sequences into lentiviral shuttle plasmid vectors The coding sequences of NLuc-tagged T1D autoantigens according to the invention were subcloned into a lentiviral shuttle plasmid upstream an IRES-GFP sequence. The lentiviral vectors were produced by calcium phosphate transient co-transfection in HEK293T cells of the plasmid containing the gene of interest sequence, the packaging plasmids (pGag-Pol and pRSV-REV), and the envelope plasmid encoding for the VSV. G protein. Medium was changed 14-16 hours after transfection and the supernatant containing the lentiviral vectors was collected 30 hours after medium change. The supernatants were filtered through a 0.22 pm filter (Millipore), transferred into sterile polyallomer tubes (Beckman Coulter) and concentrated by centrifugation at 20,000 g for 120 min at 20° C in a Beckman Optima XL-100K Ultracentrifuge. Obtained transducing units (TU) per ml titre was quantified by flow cytometry looking for GFP fluorescence after infecting HEK293T cells with serial dilutions of the collected lent...

Claims

Claims1. An in vitro method for determining the presence and / or level of at least one autoantibody in a sample, wherein the sample is a blood sample, a serum sample, or a plasma sample obtained from a human subject, the method comprising:a) contacting the sample with at least three recombinant antigens covalently linked to a bioluminescence reporter and forming at least one complex between the at least three recombinant antigens and the at least one autoantibody,b) contacting the complex formed in step a) with at least one protein selected from the group comprising or consisting of protein A, protein G, protein AG, protein L, anti IgA, anti IgM, and anti IgG; andc) determining the presence, absence, and / or amount of the at least one autoantibody in the sample that binds to the at least three recombinant antigens, wherein the at least three recombinant antigens are selected from the group consisting of: recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin and recombinant human proinsulin, andwherein the presence, absence, or amount of at least one autoantibody in the sample is determined by measuring the bioluminescence activity of the bioluminescent reporter.

2. The in vitro method according to claim 1, wherein the presence of at least one autoantibody that binds to at least one of the at least three recombinant antigens in the sample is indicative of type 1 diabetes in the human subject or of a risk of developing type 1 diabetes.

3. The in vitro method of claim 1 or 2, wherein the presence in the sample of at least two autoantibodies binding to at least two different recombinant antigens among the at least three recombinant antigens are indicative of type 1 diabetes in the human subject or of a risk of developing type 1 diabetes.

4. The in vitro method according to any one of claims 1-3, wherein the presence in the sample of at least three autoantibodies binding to the at least three recombinant antigens are indicative of type 1 diabetes in the human subject or of a risk of developing type 1 diabetes.

5. The in vitro method according to any one of claims 1-4, wherein the at least three recombinant antigens are recombinant Glutamate decarboxylase 65, recombinant human islet antigen 2 and recombinant human Zinc transporter 8.

6. The in vitro method according to any one of claims 1-5, wherein the method is performed by a luciferase immune precipitation system (LIPS) assay.

7. The in vitro method according to any one of claims 1-6, wherein the method is performed by a solid phase capture luciferase immune precipitation system (scLIPS) assay.

8. The in vitro method according to any one of claims 1-7, wherein the at least three antigens do not comprise human insulin.

9. The in vitro method according to claim 8, wherein, when at least one autoantibody is present in the sample, the sample is further contacted with recombinant human insulin.

10. The in vitro method according to any one of claims 1-9, wherein the sample is a dry sample that is reconstituted prior to the step of contacting the sample with the at least three antigens.

11. The in vitro method according to claim 10, wherein the sample is a dried blood spot (DBS).

12. A kit for detecting autoantibodies associated with type 1 diabetes, comprising:a) at least three recombinant antigens selected from the group comprising or consisting of: recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin, and recombinant human proinsulin, wherein each of the recombinant antigen molecules is fused to a bioluminescent reporter, andb) a substrate for the bioluminescent reporter.

13. The kit according to claim 12, wherein the kit comprises one or more compositions comprising the at least three antigens.

14. The kit according to claim 13, wherein the kit comprises one composition comprising the at least three antigens.

15. The kit according to anyone of claims 11-14, wherein the composition comprises recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2 and recombinant human Zinc transporter 8.

16. The kit according to anyone of claims 12-15, wherein the bioluminescent reporter is a nanoluciferase reporter protein.

17. The kit according to claim 16, wherein the nanoluciferase reporter protein is a Nanoluc luciferase (NLuc) reporter protein.

18. The kit according to claim 17, wherein the Nanoluc luciferase (NLuc) reporter protein is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99, or 100% identical to SEQ ID NO: 10.

19. The kit according to any one of claims 12-18, wherein the assay is a luciferase immune precipitation system (LIPS) assay or a solid phase capture LIPS (scLIPS) assay.

20. The kit according to claim 19, wherein the kit comprises a protein selected from the group comprising or consisting of protein A, protein G, protein AG, protein L, anti IgA, anti IgM, and anti IgG.

21. The kit according to any one of claims 12 to 20, wherein:a) the recombinant human insulin antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 1, and 11 to 46;b) the recombinant human proinsulin antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 65 to 87 and 96;c) the recombinant human islet antigen 2 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to a nucleotide sequence selected from a group consisting of SEQ ID NOs: 2 to 4;d) the recombinant human Zinc transporter 8 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 5; and / ore) the recombinant human Glutamate decarboxylase 65 antigen molecule is encoded by a nucleotide sequence at least 85, 90, 95, 98, 99 or 100% identical to the nucleotide sequence encoded by SEQ ID NO: 6.

22. A vector comprising at least one nucleic acid selected from the group consisting of: SEQ ID NOs: 1, 11 to 46, SEQ ID NOs: 65 to 87, 96, SEQ ID NOs: 2 to 4, SEQ ID NO: 5 and SEQ ID NO: 6 or a sequence at least 85, 90, 95, 98, 99 or 100% identical to said sequences.

23. A cell comprising the vector as defined in claim 22 and expressing at least one of the at least three recombinant antigens.

24. The cell according to claim 23, wherein the cell is a stably expressing transfectant cell line.

25. The cell according to claim 24, wherein the transfectant cell line is an Expi-293F cell line.

26. A method for producing the recombinant antigens of the test kit as defined in any one of claims 12 to 21, the method comprising expressing the vector as defined in claim 22 in a cell as defined in claims 23-25 and harvesting the recombinant antigen molecules.

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