Pharmaceutical preparation for inducing specific immunity against sars-cov-2

Abstract

This invention relates to biotechnology. The claimed agent can be used to prevent SARS-CoV-2. A pharmaceutical preparation has been created comprising a component (1) comprising an agent in the genomic form of a recombinant human adenovirus serotype (26) wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is disposed therein, and the pharmaceutical preparation further comprises a component (2) comprising an agent in the genomic form of a recombinant human adenovirus serotype (5) wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is disposed therein. Furthermore, a pharmaceutical formulation was created containing component 1, which comprises a drug in the genomic form of recombinant human adenovirus serotype (26) containing an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3, and the pharmaceutical formulation further contains component (2), which comprises a drug in the genomic form of recombinant simian adenovirus serotype (25) containing an expression cassette selected from SEQ ID NO: 4, SEQ ID NO: 2, and SEQ ID NO: 3. Additionally, a pharmaceutical formulation was created containing component 1, which comprises a drug in the genomic form of recombinant simian adenovirus serotype (25) containing an expression cassette selected from SEQ ID NO: 4, SEQ ID NO: 2, and SEQ ID NO: 3, and the pharmaceutical formulation further contains component (2), which comprises a drug in the genomic form of recombinant human adenovirus serotype (5) containing an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3.

Description

Technical Field

[0001] This invention relates to biotechnology, immunology, and virology. The claimed agent can be used to prevent disease caused by the severe acute respiratory syndrome virus SARS-CoV-2. Background Technology

[0002] Coronaviruses spread through person-to-person transmission via respiratory droplets, airborne dust particles, and direct contact. The estimated median incubation period is 5–6 days, followed by the appearance of initial symptoms and signs of the disease. Common symptoms of COVID-19 include fever, dry cough, shortness of breath, and fatigue. Sore throat, joint pain, runny nose, and headache have also been reported as less common symptoms.

[0003] COVID-19 can be difficult to diagnose because its symptoms are similar to those of many other viral infections. Diagnostic confirmation relies on laboratory tests that require specialized equipment, highly skilled personnel, and expensive reagents.

[0004] The clinical course of COVID-19 can range from mild to severe cases. Severe illness is more common in people over 60 years of age and in patients with chronic conditions. The most serious complications of the disease include pneumonia, acute respiratory distress syndrome, acute respiratory failure, acute heart failure, acute renal failure, septic shock, cardiomyopathy, and more. However, there are currently no specific drugs for the treatment of COVID-19.

[0005] The rapid geographical spread and high mortality rate of SARS-CoV-2 have created an urgent need to develop effective agents to prevent the disease caused by this virus. All research activities in this field are based on years of experience in product development aimed at preventing diseases caused by other members of the coronavirus family.

[0006] One solution (patent US7452542B2) proposes the use of a live attenuated vaccine to prevent disease caused by coronaviruses. This vaccine contains a live attenuated coronavirus, characterized by including a genome encoding the following: (i) tyrosine residues included in MHV-A59. 6398 Or a substituted ExoN polypeptide at a similar position, and (ii) leu included in MHV-A59 106 Orf2a peptides substituted at similar positions thereof, and pharmaceutically acceptable solvents. This invention relates to various coronaviruses, such as avian coronaviruses, animal species coronaviruses, and human coronavirus OC34.

[0007] There is a solution for obtaining a vaccine against SARS-CoV (WO2006136448A2); it involves a nucleic acid encoding an attenuated SARS-CoV virus that is capable of producing a maximum viral titer in a cell culture, said maximum viral titer being reduced by at least 2-fold compared to the maximum viral titer of wild-type SARS-CoV virus in the same cell culture.

[0008] There is a solution (RU 2 332 457C2B) for preventing coronavirus infection caused by SARS-CoV, which suggests the use of a live bacterial vaccine in which the SARS-CoV antigen is displayed on the surface of bacteria anchored with poly-γ-glutamic acid synthase (pgsBCA).

[0009] One solution (WO2016116398A1) involves the N nucleocapsid protein of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) and / or its immunogenic fragments, or nucleic acid molecules encoding the MERS-CoV N nucleocapsid protein and / or its immunogenic fragments, for use as a vaccine. This invention further discloses the use of genetic vectors selected from the group consisting of vaccinia virus, fowlpox virus, adenovirus, alpha virus, rhabdovirus, and herpesvirus for obtaining a vaccine against MERS-CoV.

[0010] One solution (WO2006071250A2) suggests using vector systems containing vaccinia virus and baculovirus containing the SARS-CoV S protein gene and its antigen fragments as a vaccine against SARS-CoV.

[0011] One solution (CN1276777C) suggests using a vaccine against severe acute respiratory syndrome based on recombinant human adenovirus serotype 5 containing the SARS-CoV virus S protein sequence.

[0012] Phylogenetic analysis showed that the SARS-CoV-2 virus is more closely related to coronaviruses found in bats (bat-SL-CoVZC45, bat-SL-CoVZXC21) than coronaviruses circulating in human populations. For example, the S protein of SARS-CoV-2 was found to have less than 75% homology with the SARS-CoV S protein (Zhou P, Yang XL, Wang XG et al. A pneumonia outbreak associated with a new coronavirus of probable bat origin. Nature. 2020;579(7798):270-273. doi:10.1038 / s41586-020-2012-7). Therefore, candidate vaccines against diseases caused by SARS-CoV are ineffective against COVID-19.

[0013] To date, there are no registered products for inducing specific immunity against the SARS-CoV-2 coronavirus. It is understood that several pharmaceutical companies are developing vaccine candidates; some of them are based on technology utilizing recombinant adenovirus vectors.

[0014] Pharmaceutical company CanSinoBIO (Tianjin, China) and the Beijing Institute of Biotechnology (Beijing, China) have jointly developed a candidate vaccine using a recombinant adenovirus type 5 vector (with the E1 and E3 regions deleted) to protect against COVID-19. This vaccine contains an optimized SARS-CoV-2 (isolated Wuhan-Hu-1) S protein gene (GenBank YP_009724390) and a tissue plasminogen activator signal peptide gene. The vaccine is formulated to contain 5 × 10⁵ molecules per 0.5 mL. 10 A liquid formulation of virus particles. This solution was chosen as a prototype by the authors of the claimed invention.

[0015] A major drawback of this solution relates to the fact that vaccines may be ineffective in some populations due to the existence of pre-existing immunity against human adenovirus type 5.

[0016] For example, according to published data, a single injection of this candidate vaccine is insufficient to induce a high level of humoral response in people aged 55 years or older (Immunogenicity and safety of a recombinant adenovirus type-5-vectored COVID-19 vaccine in healthy adults aged 18 years or older: a randomized, double-blind, placebo-controlled phase 2 trial). (trial) [Published online before print, July 20, 2020]. The Lancet. 2020; S0140-6736(20)31605-6. doi: 10.1016 / S0140-6736(20)31605-6). However, the highest risk of severe clinical course of COVID-19 is associated with retirement age.

[0017] Therefore, the background of this invention raises an urgent need to develop a pharmaceutical formulation that is safe and capable of inducing an immune response against SARS-CoV-2 coronavirus in a broad population segment. Summary of the Invention

[0018] The technical objective of the claimed inventions is to create agents for effectively inducing an immune response against the SARS-CoV-2 virus.

[0019] The technical outcome is the creation of a safe and effective pharmaceutical formulation that, through the use of two different adenovirus vectors, ensures humoral and cellular-mediated immune responses to the SARS-CoV-2 virus in diverse populations. Furthermore, the technical outcome is the creation of a pharmaceutical formulation that ensures an enhanced immune response to the SARS-CoV-2 virus.

[0020] This technical result is achieved by creating a pharmaceutical formulation for inducing specific immunity against SARS-CoV-2, the pharmaceutical formulation containing component 1, which comprises a pharmaceutical agent in the form of an expression vector based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted from the genome and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is placed, and the pharmaceutical formulation further contains component 2, which comprises a pharmaceutical agent in the form of an expression vector based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted from the genome and an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 (variant 1) is placed.

[0021] Furthermore, a pharmaceutical formulation for inducing specific immunity against SARS-CoV-2, comprising component 1, which includes an expression vector based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted from the genome and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is placed therein; and the pharmaceutical formulation further comprises component 2, which includes an expression vector based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted from the genome, wherein an expression cassette selected from SEQ ID NO: 4, SEQ ID NO: 2, and SEQ ID NO: 3 (variant 2) is placed therein.

[0022] Furthermore, a pharmaceutical formulation for inducing specific immunity against SARS-CoV-2, comprising component 1, which includes an expression vector based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted from the genome and an expression cassette selected from SEQ ID NO: 4, SEQ ID NO: 2, and SEQ ID NO: 3 is placed therein, and the pharmaceutical formulation further comprises component 2, which includes an expression vector based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted from the genome and an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 (variant 3) is placed therein.

[0023] Each pharmaceutical preparation exists in the form of a liquid preparation or a lyophilized (freeze-dried) preparation.

[0024] At this point, the buffer solution for the pharmaceutical preparation used in liquid formulations contains the following (by mass%):

[0025]

[0026] The buffer solution for pharmaceutical formulations used in lyophilized (freeze-dried) preparations contains the following (by weight%):

[0027]

[0028]

[0029] Component 1 and Component 2 are placed in different packages.

[0030] Each drug formulation is intended to induce specific immunity against the SARS-CoV-2 virus for severe acute respiratory syndrome, with components 1 and 2 administered sequentially in effective amounts, with an interval of more than one week between them. Attached Figure Description

[0031] Figure 1

[0032] The presentation box scheme is shown, in which:

[0033] 1-Promoter

[0034] 2-Target genes,

[0035] 3-Polyadecytylation signal.

[0036] Figure 2

[0037] Results of efficacy assessments of immunization using the developed drug formulation are shown, such as estimates based on the percentage of proliferative CD4+ lymphocytes restimulated by the S glycoprotein of SARS-CoV-2 on day 8 post-immunization in experimental animals.

[0038] Y-axis - Number of proliferating cells, %

[0039] X-axis - Created animal group:

[0040] 1. Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0041] 2. Ad26-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0042] 3. Ad26-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0043] 4. Ad26-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0044] 5. Ad26-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0045] 6. Ad26-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0046] 7. Ad26-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0047] 8. Ad26-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0048] 9. Ad26-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0049] 10. Ad26-null (component 1), Ad5-null (component 2);

[0050] 11. Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0051] 12. Ad26-CMV-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0052] 13. Ad26-CMV-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0053] 14. Ad26-CAG-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0054] 15. Ad26-CAG-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0055] 16. Ad26-CAG-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0056] 17. Ad26-EF1-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0057] 18. Ad26-EF1-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0058] 19. Ad26-EF1-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0059] 20. Ad26-null (component 1), simAd25-null (component 2);

[0060] 21. simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0061] 22. simAd25-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0062] 23. simAd25-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0063] 24. simAd25-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0064] 25.simAd25-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0065] 26. simAd25-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0066] 27. simAd25-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0067] 28. simAd25-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0068] 29. simAd25-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0069] 30.simAd25-null (component 1), Ad5-null (component 2);

[0070] 31. Phosphate-buffered saline solution.

[0071] ●-Data for each animal

[0072] - Geometric mean calculated for each group

[0073] Figure 3

[0074] Results of efficacy assessments of immunization with the developed drug formulation are shown, as estimated by the percentage of proliferative CD8+ lymphocytes restimulated by the S glycoprotein of the SARS-CoV-2 virus on day 8 post-immunization in mice.

[0075] Y-axis - Number of proliferating cells, %

[0076] X-axis - Created animal group:

[0077] 1. Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0078] 2. Ad26-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0079] 3. Ad26-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0080] 4. Ad26-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0081] 5. Ad26-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0082] 6. Ad26-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0083] 7. Ad26-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0084] 8. Ad26-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0085] 9. Ad26-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0086] 10. Ad26-null (component 1), Ad5-null (component 2);

[0087] 11. Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0088] 12. Ad26-CMV-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0089] 13. Ad26-CMV-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0090] 14. Ad26-CAG-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0091] 15. Ad26-CAG-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0092] 16. Ad26-CAG-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0093] 17. Ad26-EF1-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0094] 18. Ad26-EF1-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0095] 19. Ad26-EF1-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0096] 20. Ad26-null (component 1), simAd25-null (component 2);

[0097] 21. simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0098] 22. simAd25-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0099] 23. simAd25-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0100] 24. simAd25-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0101] 25.simAd25-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0102] 26. simAd25-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0103] 27. simAd25-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0104] 28. simAd25-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0105] 29. simAd25-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0106] 30.simAd25-null (component 1), Ad5-null (component 2);

[0107] 31. Phosphate-buffered saline solution.

[0108] ●-Data for each animal shown

[0109] - Geometric mean calculated for each group

[0110] Figure 4

[0111] Survival curves of golden Syrian hamsters immunized with the developed drug formulation and control group using a lethal SARS-CoV-2 virus infection model are shown.

[0112] Y-axis - Animal survival rate, %

[0113] X-axis - Number of days after attack with the SARS-CoV-2 virus.

[0114] ●- Presents the survival rate of golden Syrian hamsters immunized with the developed drug formulation, in the created group:

[0115] 1) Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0116] 2) Ad26-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0117] 3) Ad26-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0118] 4) Ad26-CAG-S-CoV2 component 1), Ad5-CMV-S-CoV2 component 2;

[0119] 5) Ad26-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0120] 6) Ad26-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0121] 7) Ad26-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0122] 8) Ad26-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0123] 9) Ad26-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0124] 11) Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0125] 12) Ad26-CMV-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0126] 13) Ad26-CMV-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0127] 14) Ad26-CAG-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0128] 15) Ad26-CAG-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0129] 16) Ad26-CAG-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0130] 17) Ad26-EF1-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0131] 18) Ad26-EF1-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0132] 19) Ad26-EF1-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0133] 21) simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0134] 22) simAd25-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0135] 23) simAd25-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0136] 24) simAd25-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0137] 25) simAd25-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0138] 26) simAd25-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0139] 27) simAd25-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0140] 28) simAd25-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0141] 29) simAd25-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2),

[0142] It reached 100% in all groups.

[0143] ◆-Negative control group:

[0144] 10) Ad26-null (component 1), Ad5-null (component 2).

[0145] ■-Negative control group:

[0146] 20)Ad26-null (component 1), simAd25-null (component 2).

[0147] -Negative control group:

[0148] 30)simAd25-null (component 1), Ad5-null (component 2).

[0149] ▲- Negative control group:

[0150] 32.32)-phosphate buffered saline.

[0151] Figure 5

[0152] The results of an assessment of humoral immune responses to SARS-CoV2 viral antigens in primates immunized with a drug formulation developed according to variant 1 are presented.

[0153] Y-axis - IgG titers against SARS-CoV-2 RBD.

[0154] X-axis - number of days.

[0155] ●-Immunize with a drug formulation developed based on variant 1 (Ad26-CMV-S-SARS-CoV-2; Ad5-CMV-S-SARS-CoV-2).

[0156] ○ - Placebo.

[0157] Figure 6

[0158] Results of efficacy assessments of immunization using the developed drug formulation are shown, as estimated by the percentage of proliferative CD4+ lymphocytes restimulated by the RBD fragment of the SARS-CoV-2S antigen on day 8 post-immunization in primates.

[0159] Y-axis - Number of proliferating cells, %

[0160] X-axis - number of days.

[0161] ЧepньIм цветом oбo3HaчeHa Гpyппa жHBOTHbIX, HMMyHH3HpoBaHHЪIxpa3paбoTaHHЪIM φapMaцeBTичecκHM cpeдcTBOM ∏o BapиaHTy 1(Ad26-CMV-S-SARS-CoV-2; Ad5-CMV-S-SARS-CoV-2).

[0162] Black was used to describe the animal group immunized with a drug formulation developed based on variant 1 (Ad26-CMV-S-SARS-CoV-2; Ad5-CMV-S-SARS-CoV-2).

[0163] The control group (unvaccinated animals) is shown in gray.

[0164] The arithmetic mean is shown as a dashed line for each data group. Statistically significant differences between values ​​obtained from immunized animals and control (unvaccinated) animals are indicated in parentheses and asterisks (Mann-Whitney test, p < 0.05).

[0165] Figure 7

[0166] Results of efficacy assessments of immunization using the developed drug formulation are shown, such as estimates based on the percentage of proliferating CD8+ lymphocytes restimulated by the RBD fragment of the SARS-CoV-2S antigen on day 8 post-immunization in primates.

[0167] Y-axis - Number of proliferating cells, %

[0168] X-axis - number of days.

[0169] Black was used to describe the animal group immunized with a drug formulation developed based on variant 1 (Ad26-CMV-S-SARS-CoV-2; Ad5-CMV-S-SARS-CoV-2).

[0170] The control group (unvaccinated animals) is shown in gray.

[0171] The arithmetic mean is shown as a dashed line for each data group. Statistically significant differences between values ​​obtained from immunized animals and control (unvaccinated) animals are indicated in parentheses and by the symbol *, with p < 0.05 (Mann-Whitney test).

[0172] Figure 8

[0173] Results of efficacy assessments of immunization in volunteers using a liquid formulation of the drug developed according to variant 1 are shown, as estimated by the percentage of proliferating CD8+ lymphocytes restimulated by the SARS-CoV-2S antigen.

[0174] Y-axis - Number of proliferating cells, %

[0175] X-axis - number of days.

[0176] ● - A symbol used for each volunteer on day 0.

[0177] ■ - A symbol used for each volunteer on day 14.

[0178] ▲ - The symbol used for each volunteer on day 28.

[0179] For each data set, the median is shown as a black line. Statistically significant differences between values ​​obtained on day 0, day 14, and day 28 are indicated in parentheses and the symbols *, p < 0.05; **, p < 0.01; ****, p < 0.001 (Mann-Whitney test).

[0180] Figure 9

[0181] Results of efficacy assessments of immunization in volunteers using a liquid formulation of the drug developed according to variant 1 are shown, as estimated by the percentage of proliferating CD4+ lymphocytes restimulated by the SARS-CoV-2S antigen.

[0182] Y-axis - Number of proliferating cells, %

[0183] X-axis - number of days.

[0184] ● - A symbol used for each volunteer on day 0.

[0185] ■ - A symbol used for each volunteer on day 14.

[0186] ▲ - The symbol used for each volunteer on day 28.

[0187] For each data set, the median is shown as a black line. Statistically significant differences between values ​​obtained on day 0, day 14, and day 28 are indicated in parentheses and the symbols *, p < 0.05; **, p < 0.01; ****, p < 0.001 (Mann-Whitney test).

[0188] Figure 10

[0189] Results of an evaluation of the effectiveness of immunization in volunteers using a lyophilized (freeze-dried) formulation of a drug formulation developed according to variant 1 are shown, as estimated by the percentage of proliferating CD8+ lymphocytes restimulated by the SARS-CoV-2S antigen.

[0190] Y-axis - Number of proliferating cells, %

[0191] X-axis - number of days.

[0192] ● - A symbol used for each volunteer on day 0.

[0193] ■ - A symbol used for each volunteer on day 14.

[0194] ▲ - The symbol used for each volunteer on day 28.

[0195] For each data set, the median is shown as a black line. Statistically significant differences between values ​​obtained on day 0, day 14, and day 28 are indicated in parentheses and the symbols *, p < 0.05; **, p < 0.01; ****, p < 0.001 (Mann-Whitney test).

[0196] Figure 11

[0197] Results of an evaluation of the effectiveness of immunization in volunteers using a lyophilized (freeze-dried) formulation of a drug formulation developed according to variant 1 are shown, as estimated by the percentage of proliferating CD4+ lymphocytes restimulated by the SARS-CoV-2S antigen.

[0198] Y-axis - Number of proliferating cells, %

[0199] X-axis - number of days.

[0200] ● - A symbol used for each volunteer on day 0.

[0201] ■ - A symbol used for each volunteer on day 14.

[0202] ▲ - The symbol used for each volunteer on day 28.

[0203] For each data set, the median is shown as a black line. Statistically significant differences between values ​​obtained on day 0, day 14, and day 28 are indicated in parentheses and the symbols *, p < 0.05; **, p < 0.01; ****, p < 0.001 (Mann-Whitney test).

[0204] Figure 12

[0205] The study showed an increase (fold) in the concentration of IFNγ in the culture medium of peripheral blood mononuclear cells from volunteers immunized with a liquid formulation of the drug developed according to variant 1, before immunization (day 0) and on days 14 and 28 of the study, after they were restimulated by the SARS-CoV-2S antigen.

[0206] The Y-axis IFN-γ concentration increased (by a factor of 1).

[0207] X-axis - number of days.

[0208] ● - A symbol used to display the value obtained by each volunteer on day 0.

[0209] ■ - A symbol used to display the value obtained by each volunteer on day 14.

[0210] ▲ - A symbol used to show the value obtained by each volunteer on day 28.

[0211] For each data set, the median is shown as a black line. Statistically significant differences between values ​​obtained on day 0, day 14, and day 28 are indicated in parentheses and by the symbol *, p < 0.05; ****, p < 0.001 (Mann-Whitney test).

[0212] Figure 13

[0213] The study showed an increase (fold) in the concentration of IFNγ in the culture medium of peripheral blood mononuclear cells from volunteers immunized with a lyophilized (freeze-dried) formulation of a drug formulation developed according to variant 1, before immunization (day 0) and on days 14 and 28 of the study, after they were restimulated by the SARS-CoV-2S antigen.

[0214] The Y-axis IFN-γ concentration increased (by a factor of 1).

[0215] X-axis - number of days.

[0216] • - A symbol used to display the value obtained by each volunteer on day 0.

[0217] ■ - A symbol used to display the value obtained by each volunteer on day 14.

[0218] ▲ - A symbol used to show the value obtained by each volunteer on day 28.

[0219] The dots depict the values ​​for each volunteer participating in the study. For each data group, the median is shown as a black line. Statistically significant differences between values ​​obtained on day 0, day 14, and day 28 are indicated in parentheses and by the symbols *, p < 0.05; ****, p < 0.001 (Mann-Whitney test).

[0220] Figure 14

[0221] The results of an assessment of humoral immune responses to SARS-CoV2 viral antigens in volunteers immunized with a liquid formulation of a drug formulation developed according to variant 1 are shown.

[0222] Y-axis - IgG titer against SARS-CoV-2S glycoprotein RBD.

[0223] X-axis - number of days.

[0224] - Data for each volunteer.

[0225] Figure 15

[0226] The results of an assessment of humoral immune responses to SARS-CoV2 viral antigens in volunteers immunized with a lyophilized (freeze-dried) formulation of a drug formulation developed according to variant 1 are shown.

[0227] Y-axis - IgG titer against SARS-CoV-2S glycoprotein RBD.

[0228] X-axis - number of days.

[0229] - Data for each volunteer. Detailed Implementation

[0230] To create a safe and effective drug formulation for inducing a specific immune response against the SARS-CoV-2 virus, an adenovirus vector system was chosen. Adenovirus vectors are characterized by many advantages, including: inability to replicate in human cells; the possibility of entering both dividing and non-dividing human cells; the ability to induce both cell-mediated and humoral immune responses; and the potential to ensure high-level expression of the target antigen.

[0231] Meanwhile, the clinical application of these vectors may be limited because some individuals with a history of adenovirus infection may have a pre-existing immune response to adenovirus. Research has shown that antibody titers against adenovirus vectors increase with age and vary across different population segments. In this context, high seropositivity rates for human adenovirus serotype 5 have been reported in 40–45% of the US population and up to 90% of the sub-Saharan African population. (Prevalence of neutralizing antibodies against adenovirus serotypes 5 and 35 in the adult populations of The Gambia, South Africa, and the United States. Clin. Diagn. Lab. Immunol. 2004; 11(2): 351-357; Dudareva M, Andrews L, Gilbert SC et al. Prevalence of serum neutralizing antibodies against chimpanzeeadenovirus 63 and human adenovirus 5 in Kenyan children, in the context of vaccine vector efficacy.) efficacy). Vaccine. 2009;27(27):3501-3504; S. Zhang S, Huang W, Zhou X, Zhao Q, Wang Q, Jia B. Seropervalence of neutralizing antibodies to human adenoviruses type-5 and type-26 and chimpanzee adenovirus type-68 in healthy Chinese adults. Journal of Medical Virology. 2013;85(6):1077-1084.

[0232] Neutralizing antibodies against the vector lead to a significant reduction in the specific immune response to the transgene and may reduce the effectiveness of immunization.

[0233] Based on the research conducted, the inventors identified adenovirus vector serotypes with such genetic differences that would exclude any influence on the generation of antigen-specific immune responses against vaccine antigens during sequential immunization.

[0234] Three viruses were selected for further research—human adenovirus serotype 26, human adenovirus serotype 5, and simian adenovirus serotype 25. In the next phase, viral clones differentiated by superior growth kinetics were selected. These clones were used to create genetically engineered recombinant adenovirus vectors.

[0235] Therefore, the combination of several types of genetic vectors utilized has supported the development of a range of pharmaceutical formulations to overcome difficulties associated with pre-existing human immune responses to some adenoviruses, particularly human adenovirus serotype 5.

[0236] Therefore, the present invention can be used in which a variant of the pharmaceutical formulation is selected after assessing the patient’s immunity to the adenovirus vector serotype (human adenovirus serotype 26, human adenovirus serotype 5, simian adenovirus serotype 25) contained in the pharmaceutical formulation.

[0237] Using genetic engineering techniques, expression cassettes were placed within recombinant adenovirus vectors. These cassettes contain vaccine antigen genes and expression regulatory elements (promoters and polyadenylation signals). A schematic diagram of the expression cassettes is shown below. Figure 1 As shown above.

[0238] To maximize the effectiveness of inducing an immune response, the authors requested the protection of multiple variants of the expression cassette.

[0239] The spike (S) protein of the SARS-CoV-2 virus, optimized for expression in mammalian cells, was used as the antigen in all boxes. The S protein is one of the structural proteins of the coronavirus. It is exposed on the surface of the viral particle and is responsible for binding the virus to the ACE2 (angiotensin-converting enzyme 2) receptor. Results from completed studies indicate the generation of virus-neutralizing antibodies against the S protein, thus suggesting it as a promising antigen for the development of drug formulations.

[0240] The expression cassette SEQ ID NO: 1 contains the CMV promoter, the SARS-CoV-2 viral S protein gene, and a polyadenylation signal. The CMV promoter is the promoter of the immediate early genes of cytomegalovirus, ensuring constitutive expression in a variety of cell types. However, the expression intensity of the target genes controlled by the CMV promoter varies with different cell types. Furthermore, the level of transgene expression under the control of the CMV promoter shows a decrease with increasing cell culture duration. This is due to the repression of gene expression associated with DNA methylation [Wang W., Jia YL., Li YC., Jing CQ., Guo X., Shang XF., Zhao CP., Wang TY. Impact of different promoters, promoter mutation, and an enhancer recombinant protein expression in CHO cells. / / Scientific Reports - 2017 - Vol. 8 - p. 10416].

[0241] The expression cassette SEQ ID NO: 2 contains the CAG promoter, the SARS-CoV-2 viral S protein gene, and a polyadenylation signal. The CAG promoter is a synthetic promoter containing an early enhancer of the CMV promoter, a chicken β-actin promoter, and chimeric introns (chicken β-actin and rabbit β-globin). Experiments have shown that the CAG promoter has higher transcriptional activity compared to the CMV promoter [Evaluation of three different promoters driving gene expression in developing chicken embryo by using in vivo electroporation. / / Genet.Mol.Res.-2014-Vol.13-pp.1270-1277].

[0242] The expression cassette SEQ ID NO: 3 contains the EF1 promoter, the SARS-CoV-2 viral S protein gene, and a polyadenylation signal. The EF1 promoter is the promoter of human eukaryotic translation elongation factor 1α (EF-1α). This promoter is constitutively active in a variety of cell types [PMID: 28557288. The EF-1α promoter maintains high-level transgene expression episomal vectors in transfected CHO-K1 cells]. The EF-1α gene encodes elongation factor 1α, one of the most common proteins in eukaryotic cells and is expressed in almost all mammalian cell types. The EF-1α promoter frequently exhibits activity in cells where viral promoters fail to promote the expression of the controlled gene and in cells where viral promoters are gradually eliminated.

[0243] The expression cassette SEQ ID NO:4 contains the CMV promoter, the SARS-CoV-2 viral S protein gene, and a polyadenylation signal.

[0244] Therefore, as a result of the completed task, the following three variants of the pharmaceutical formulation were developed.

[0245] 1. A pharmaceutical preparation for inducing specific immunity against SARS-CoV-2, the pharmaceutical preparation comprising component 1, the component comprising an expression vector of the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted from the genome and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is disposed therein, and the pharmaceutical preparation further comprising component 2, the component comprising an expression vector of the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted from the genome, wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is disposed therein.

[0246] 2. A pharmaceutical preparation for inducing specific immunity against SARS-CoV-2, the pharmaceutical preparation comprising component 1, the component comprising an expression vector of the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted from the genome and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is placed therein, and the pharmaceutical preparation further comprising component 2, the component comprising an expression vector of the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted from the genome, wherein an expression cassette selected from SEQ ID NO: 4, SEQ ID NO: 2, and SEQ ID NO: 3 is placed therein.

[0247] 3. A pharmaceutical preparation for inducing specific immunity against SARS-CoV-2, the pharmaceutical preparation comprising component 1, the component comprising an expression vector of the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted from the genome and an expression cassette selected from SEQ ID NO: 4, SEQ ID NO: 2, and SEQ ID NO: 3 is placed therein, and the pharmaceutical preparation further comprising component 2, the component comprising an expression vector of the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted from the genome and an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is placed therein.

[0248] In this way, the components of a pharmaceutical preparation can be placed in different packages.

[0249] In addition, the authors of this invention have developed liquid formulations and lyophilized (freeze-dried) formulations of pharmaceutical preparations.

[0250] In addition, the inventors chose such a variant of the buffer solution, which allows the developed pharmaceutical formulation to be stored at temperatures below -18°C and freeze-dried (lyophilized) in a temperature range of +2°C to +8°C.

[0251] In addition, a method for using the drug formulation to induce specific immunity against the SARS-CoV-2 virus for severe acute respiratory syndrome has been developed, wherein components 1 and 2 are administered in effective amounts sequentially, with an interval of more than one week.

[0252] The implementation of this invention is demonstrated by the following examples:

[0253] Example 1.

[0254] Preparation of expression vectors containing the genome of recombinant human adenovirus serotype 26.

[0255] In the first phase, the design of the plasmid construct pAd26-Ends was proposed. It carries two regions (two homologous arms) homologous to the genome of recombinant human adenovirus serotype 26 and an ampicillin resistance gene. One homologous arm is the start portion of the recombinant human adenovirus serotype 26 genome (from the left inverted terminal repeat sequence to the E1 region and the viral genome sequence containing the pIX protein). The other homologous arm contains a nucleotide sequence located after the ORF3 E4 region up to the end of the genome. The synthesis of the pAd26-Ends construct was performed by the Moscow company "Eurogen" ZAO.

[0256] Human adenovirus serotype 26 DNA isolated from the virion was mixed with pAd26-Ends. The plasmid pAd26-dlE1, carrying the genome of human adenovirus serotype 26 with a deleted E1 region, was obtained through homologous recombination between pAd26-Ends and viral DNA.

[0257] Then, in the obtained plasmid pAd26-dlE1, using standard cloning techniques, the sequence containing open reading frame 6 (ORF6-Ad26) was replaced with a similar sequence from the genome of human adenovirus serotype 5. This operation aimed to ensure efficient replication of human adenovirus serotype 26 in HEK293 cell cultures. As a result, the plasmid pAd26-dlE1-ORF6-Ad5 was derived.

[0258] Furthermore, using standard genetic engineering techniques, the E3 region of the adenovirus genome (approximately 3321 base pairs between gene pVIII and exon U) was deleted from the constructed plasmid pAd26-dlE1-ORF6-Ad5 to expand the vector's packaging capacity. Ultimately, the recombinant vector pAd26-only-null, based on the genome of human adenovirus serotype 26, was obtained, possessing the open reading frame ORF6 of human adenovirus serotype 5 and having the deleted E1 and E3 regions. Sequence SEQ ID NO: 5 was used as the parental sequence for human adenovirus serotype 26.

[0259] In addition, the author developed several designs for the expression box:

[0260] - Expression cassette SEQ ID NO: 1 contains the CMV promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal;

[0261] - Expression cassette SEQ ID NO: 2 contains the CAG promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal;

[0262] - Expression cassette SEQ ID NO:3 contains the EF1 promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal.

[0263] Based on the plasmid construct pAd26-Ends, the constructs pArms-26-CMV-S-CoV2, pArms-26-CAG-S-CoV2, and pArms-26-EF1-S-CoV2 were obtained using genetic engineering techniques. These constructs contain expression cassettes SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3, respectively, and a homologous arm carrying the genome of human adenovirus serotype 26.

[0264] Next, the constructs pArms-26-CMV-S-CoV2, pArms-26-CAG-S-CoV2, and pArms-26-EF1-S-CoV2 were linearized through unique hydrolysis sites between homologous arms; each plasmid was then mixed with the recombinant vector pAd26-only-null.

[0265] Homologous recombination allows the acquisition of plasmids pAd26-only-CMV-S-CoV2, pAd26-only-CAG-S-CoV2, and pAd26-only-EF1-S-CoV2, which carry the genome of recombinant human adenovirus serotype 26 and have deletions of the open reading frame ORF6 and E1 and E3 regions of human adenovirus serotype 5, and have expression cassettes SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3, respectively.

[0266] During the fourth phase, plasmids pAd26-only-CMV-S-CoV2, pAd26-only-CAG-S-CoV2, and pAd26-only-EF1-S-CoV2 were hydrolyzed with specific restriction endonucleases to remove the vector portion. The derived DNA products were used to transfect HEK293 cell cultures.

[0267] Therefore, an expression vector containing the genome of recombinant human adenovirus serotype 26 was obtained, wherein the E1 and E3 regions were deleted and the RF6-Ad26 region was replaced by ORF6-Ad5, wherein expression cassettes selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 were integrated.

[0268] Example 2.

[0269] An immunobiological agent in the form of an expression vector based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is integrated.

[0270] At this stage, anion exchange and size exclusion chromatography were used to purify the expression vector obtained in Example 1. The final suspension contained adenovirus particles in a buffer solution for a liquid formulation of a pharmaceutical preparation or in a buffer solution for a lyophilized (freeze-dried) formulation of a pharmaceutical preparation.

[0271] Therefore, the following immunobiological agent was produced based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions were deleted and the ORF6-Ad26 region was replaced by ORF6-Ad5:

[0272] 1. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein the expression cassette SEQ ID NO: 1 (Ad26-CMV-S-CoV2) containing the CMV promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0273] 2. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein the expression cassette SEQ ID NO: 1 (Ad26-CMV-S-CoV2) containing the CMV promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0274] 3. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein the expression cassette SEQ ID NO: 2 (Ad26-CAG-S-CoV2) containing the CAG promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0275] 4. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein the expression cassette SEQ ID NO: 2 (Ad26-CAG-S-CoV2) containing the CAG promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0276] 5. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein the expression cassette SEQ ID NO: 3 (Ad26-EF1-S-CoV2) containing the EF1 promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0277] 6. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein the expression cassette SEQ ID NO: 3 (Ad26-EF1-S-CoV2) containing the EF1 promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0278] Each presented immunobiological agent is component 1 in variant 1 and variant 2 of the developed drug formulation.

[0279] Example 3.

[0280] Preparation of expression vectors containing the genome of recombinant simian adenovirus serotype 25.

[0281] In the first phase, the design of the plasmid construct pSim25-Ends was proposed. It carries two regions (two homologous arms) homologous to the genome of simian adenovirus serotype 25. One homologous arm contains the start portion of the simian adenovirus serotype 25 genome (from the left inverted terminal repeat sequence to the E1 region) and the sequence from the end of the E1 region to the pIVa2 protein. The other homologous arm contains the terminal sequence of the adenovirus genome, including the right inverted terminal repeat sequence. The synthesis of the pSim25-Ends construct was performed by the Moscow-based company “Eurogen” ZAO.

[0282] The DNA of simian adenovirus serotype 25 isolated from the virion was mixed with pSim25-Ends. Through homologous recombination between pSim25-Ends and viral DNA, plasmid pSim25-dlE1 was obtained, which carries the genome of simian adenovirus serotype 25 with the E1 region deleted.

[0283] Furthermore, using standard genetic engineering techniques, the E3 region of the adenovirus genome (approximately 3921 base pairs from the start of gene 12,5K to gene 14.7K) was deleted from the constructed plasmid pSim25-dlE1 to expand the vector's packaging capacity. Finally, the plasmid construct pSim25-null was obtained, encoding the complete genome of simian adenovirus serotype 25 with the deleted E1 and E3 regions. Sequence SEQ ID NO: 6 was used as the parental sequence for simian adenovirus serotype 25.

[0284] In addition, the author developed several designs for the expression box:

[0285] - Expression cassette SEQ ID NO:4 contains the CMV promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal;

[0286] - Expression cassette SEQ ID NO: 2 contains the CAG promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal;

[0287] - Expression cassette SEQ ID NO:3 contains the EF1 promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal.

[0288] Then, based on the plasmid construct pSim25-Ends, genetic engineering techniques were used to obtain the constructs pArms-Sim25-CMV-S-CoV2, pArms-Sim25-CAG-S-CoV2, and pArms-Sim25-EF1-S-CoV2. These subsequent constructs each contain an expression cassette (SEQ ID NO: 4, SEQ ID NO: 2, or SEQ ID NO: 3) and a homologous arm carrying the genome from simian adenovirus serotype 25. Next, the constructs pArms-Sim25-CMV-S-CoV2, pArms-Sim25-CAG-S-CoV2, and pArms-Sim25-EF1-S-CoV2 were linearized through unique hydrolysis sites between the homologous arms; each plasmid was then mixed with the recombinant vector pSim25-null. Homologous recombination allows the acquisition of plasmid vectors pSim25-CMV-S-CoV2, pSim25-CAG-S-CoV2, and pSim25-EFl-S-CoV2, which respectively contain the complete genome of recombinant human adenovirus serotype 26 (with an open reading frame ORF6 containing simian adenovirus serotype 25 (with deleted E1 and E3 regions)) and expression cassettes SEQ ID NO: 4, SEQ ID NO: 2, or SEQ ID NO: 3.

[0289] During the third phase, plasmids pSim25-CMV-S-CoV2, pSim25-CAG-S-CoV2, and pSim25-EF1-S-CoV2 were hydrolyzed with specific restriction endonucleases to remove the vector portion. The derived DNA products were used to transfect HEK293 cell cultures. The resulting material was used to generate prepared quantities of recombinant adenovirus.

[0290] As a result, recombinant human adenovirus serotype 25 containing the SARS-CoV-2 virus S protein gene was obtained; simAd25-CMV-S-CoV2 (containing expression cassette SEQ ID NO: 2) and simAd25-EF1-S-CoV2 (containing expression cassette SEQ ID NO: 3).

[0291] Therefore, an expression vector containing the genome of recombinant simian adenovirus 25 was obtained, wherein the E1 and E3 regions were deleted, and expression cassettes selected from SEQ ID NO: 4, SEQ ID NO: 2, and SEQ ID NO: 3 were integrated.

[0292] Example 4.

[0293] Production of an immunobiological agent in the form of an expression vector based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted, and an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is integrated.

[0294] At this stage, the expression vector obtained in Example 3 was purified using anion exchange and size exclusion chromatography. The final suspension contained adenovirus particles in either a buffer solution for a liquid formulation of a pharmaceutical preparation or a buffer solution for a lyophilized (freeze-dried) formulation of a pharmaceutical preparation.

[0295] Therefore, the following immunobiological agents were produced based on the genome of simian adenovirus serotype 25, in which the E1 and E3 regions were deleted:

[0296] 1. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 1 (simAd25-CMV-S-CoV2) containing the CMV promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0297] 2. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 1 (simAd25-CMV-S-CoV2) containing the CMV promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0298] 3. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 2 (simAd25-CAG-S-CoV2) containing the CAG promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0299] 4. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 2 (simAd25-CAG-S-CoV2) containing the CAG promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0300] 5. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 3 (simAd25-EF1-S-CoV2) containing the EF1 promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0301] 6. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 3 (simAd25-EF1-S-CoV2) containing the EF1 promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0302] Each presented immunobiological agent includes component 2 in variant 1 of the developed drug formulation and component 1 in variant 3 of the developed drug formulation.

[0303] Example 5.

[0304] Preparation of expression vectors containing the genome of recombinant human adenovirus serotype 5.

[0305] In the first phase, the design of the plasmid construct pAd5-Ends was proposed. It carries two regions (two homologous arms) homologous to the genome of recombinant human adenovirus serotype 5. One homologous arm contains the start portion of the recombinant human adenovirus serotype 5 genome (from the left inverted terminal repeat to the E1 region) and the sequence of the viral genome containing the pIX protein. The other homologous arm contains a nucleotide sequence following the ORF3 E4 region up to the end of the genome. The synthesis of the pAd26-Ends construct was performed by the Moscow-based company “Eurogen” ZAO.

[0306] Human adenovirus serotype 5 DNA isolated from the virion was mixed with pAd26-Ends. Through homologous recombination between pAd26-Ends and viral DNA, plasmid pAd26-dlE1 was obtained, which carries the genome of human adenovirus serotype 5 with the E1 region deleted.

[0307] Furthermore, using standard genetic engineering techniques, the E3 region of the adenovirus genome (2685 base pairs from the end of gene 12.5K to the start of the U-exon sequence) was deleted from the constructed plasmid pAd5-dlE1 to expand the vector's packaging capacity. Ultimately, the recombinant plasmid vector pAd5-too-null, based on the genome of human adenovirus serotype 5 and lacking both the E1 and E3 regions, was obtained. The sequence SEQ ID NO: 7 was used as the parental sequence for human adenovirus serotype 5.

[0308] In addition, the author developed several designs for the expression box:

[0309] - Expression cassette SEQ ID NO: 1 contains the CMV promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal;

[0310] - Expression cassette SEQ ID NO: 2 contains the CAG promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal;

[0311] - Expression cassette SEQ ID NO:3 contains the EF1 promoter, the SARS-CoV-2 virus S protein gene, and a polyadenylation signal.

[0312] Then, based on the plasmid construct pAd5-Ends, pArms-Ad5-CMV-S-CoV2, pArms-Ad5-CAG-S-CoV2, and pArms-Ad5-EF1-S-CoV2 were obtained using genetic engineering techniques. These subsequent constructs contain expression cassettes SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3, respectively, and a homologous arm carrying the genome of human adenovirus serotype 5.

[0313] Next, the constructs pArms-Ad5-CMV-S-CoV2, pArms-Ad5-CAG-S-CoV2, and pArms-Ad5-EF1-S-CoV2 were linearized via unique hydrolysis sites between homologous arms; each plasmid was then mixed with the recombinant vector pAd5-too-null. Homologous recombination allowed the acquisition of plasmids pAd5-too-CMV-S-CoV2, pAd5-too-GAC-S-CoV2, and pAd5-too-EF1-S-CoV2, which carried the genome of recombinant human adenovirus serotype 5 with the E1 and E3 regions deleted, respectively, and expression cassettes SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3.

[0314] During the fourth phase, plasmids pAd5-too-CMV-S-CoV2, pAd5-too-GAC-S-CoV2, and pAd5-too-EF1-S-CoV2 were hydrolyzed with specific restriction endonucleases to remove the vector portion. The derived DNA products were used to transfect HEK293 cell cultures. The resulting material was used to accumulate prepared quantities of recombinant adenovirus.

[0315] As a result, recombinant human adenovirus serotype 5 was obtained, which contains the SARS-CoV-2 virus S protein gene; Ad5-CMV-S-CoV2 (containing expression cassette SEQ ID NO: 1), Ad5-CAG-S-CoV2 (containing expression cassette SEQ ID NO: 2), and Ad5-EF1-S-CoV2 (containing expression cassette SEQ ID NO: 3).

[0316] Therefore, an expression vector containing the genome of recombinant human adenovirus 5 was obtained, wherein the E1 and E3 regions were deleted, and expression cassettes selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 were integrated.

[0317] Example 6.

[0318] Production of an immunobiological agent in the form of an expression vector based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted, and an expression cassette selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 is integrated.

[0319] At this stage, the expression vector obtained in Example 5 was purified using anion exchange and size exclusion chromatography. The final suspension contained adenovirus particles in either a buffer solution for a liquid formulation of a pharmaceutical preparation or a buffer solution for a lyophilized (freeze-dried) formulation of a pharmaceutical preparation.

[0320] Therefore, the following immunobiological agents were produced based on the genome of recombinant human adenovirus serotype 5, in which the E1 and E3 regions were deleted:

[0321] 1. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 1 (Ad5-CMV-S-CoV2) containing the CMV promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0322] 2. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 1 (Ad5-CMV-S-CoV2) containing the CMV promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0323] 3. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted, and an expression cassette containing the CAG promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal SEQ ID NO: 2 (Ad5-CAG-S-CoV2) in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0324] 4. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 2 (Ad5-CAG-S-CoV2) containing the CAG promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0325] 5. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted, and an expression cassette containing the EF1 promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal, SEQ ID NO: 3 (Ad5-EF1-S-CoV2), is in a buffer solution of a liquid formulation for pharmaceutical preparation.

[0326] 6. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, wherein the E1 and E3 regions are deleted, wherein the expression cassette SEQ ID NO: 3 (Ad5-EF1-S-CoV2) containing the EF1 promoter, the SARS-CoV-2 virus S protein gene and the polyadenylation signal is in a buffer solution of a lyophilized (freeze-dried) formulation for use in pharmaceutical preparations.

[0327] Each presented immunobiological agent includes component 1 from variant 1 and variant 2 of the developed drug formulation.

[0328] Each presented immunobiological agent includes component 2 from variant 1 and variant 3 of the developed drug formulation.

[0329] Example 7.

[0330] Production of buffer solutions

[0331] The pharmaceutical formulation developed according to the present invention consists of two components placed in different vials. Thus, each component comprises, in a buffer solution, an immunobiological agent based on a recombinant adenovirus having an expression cassette.

[0332] The inventors have selected the composition of the buffer solution to ensure the stability of the recombinant virus particles. The solution contains:

[0333] 1. Tris(hydroxymethyl)aminomethane (Tris) required to maintain the pH of the solution.

[0334] 2. Add sodium chloride to achieve the necessary ionic forces and osmotic pressure.

[0335] 3. Sucrose used as a cryoprotectant.

[0336] 4. Magnesium chloride hexahydrate is required as a source of divalent cations.

[0337] 5. EDTA as an inhibitor of free radical oxidation.

[0338] 6. Polysorbate-80, used as a source of surfactant.

[0339] 7. 95% ethanol as an inhibitor of free radical oxidation.

[0340] 8. Water used as a solvent.

[0341] The authors of this invention have developed two variants of the buffer solution: a liquid formulation for pharmaceutical preparations and a lyophilized (freeze-dried) formulation for pharmaceutical preparations.

[0342] To estimate the concentrations of substances contained in the buffer solutions of liquid formulations used in pharmaceutical preparations, several options for the experimental group were developed (Table 1). One of the components of the pharmaceutical preparation was added to each produced buffer solution:

[0343] 1. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, containing an expression cassette with a CMV promoter, SARS-CoV-2 viral S protein gene, and adenosine monophosphate signal, 1*10 11 A virus particle.

[0344] 2. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, containing an expression cassette with a CMV promoter, SARS-CoV-2 viral S protein gene, and adenosine monophosphate signal, 1*102 11 A virus particle.

[0345] 3. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, containing an expression cassette with the CMV promoter, SARS-CoV-2 viral S protein gene, and adenosine monophosphate signal, 1*10 11 A virus particle.

[0346] Therefore, the stability of each adenovirus serotype included in the drug formulation was confirmed. The obtained drug formulations were stored at -18°C and -70°C for 3 months, then thawed; and changes in the titer of the recombinant adenovirus were assessed.

[0347] Table 1 - Composition of experimental buffer solutions for liquid formulations of pharmaceutical preparations

[0348] Table 1.

[0349]

[0350] The results of the experiments showed that the titer of recombinant adenovirus did not change after 3 months of storage in a buffer solution of liquid formulation for pharmaceutical preparations at temperatures of -18°C and -70°C.

[0351] Therefore, the buffer solution developed for liquid formulations of pharmaceutical preparations ensures the stability of all components of the developed pharmaceutical preparations within the following active fraction (mass%) range:

[0352] Tris: 0.1831% by mass to 0.3432% by mass;

[0353] Sodium chloride: 0.3313% by mass to 0.6212% by mass;

[0354] Sucrose: 3.7821% by mass to 7.0915% by mass;

[0355] Magnesium chloride hexahydrate: 0.0154% by mass to 0.0289% by mass;

[0356] EDTA: 0.0029% by mass to 0.0054% by mass;

[0357] Polysorbate-80: 0.0378% by mass to 0.0709% by mass;

[0358] 95% ethanol: 0.0004% by mass to 0.0007% by mass;

[0359] Solvent: The remainder.

[0360] To estimate the concentrations of substances contained in the buffer solutions of lyophilized (freeze-dried) formulations used in pharmaceutical preparations, several options for the experimental group were proposed (Table 2). One component of the pharmaceutical preparation was added to the buffer solution of each production:

[0361] 1. An immunobiological agent based on the genome of recombinant human adenovirus serotype 26, containing an expression cassette with a CMV promoter, SARS-CoV-2 viral S protein gene, and adenosine monophosphate signal, 1*10 11 A virus particle.

[0362] 2. An immunobiological agent based on the genome of recombinant human adenovirus serotype 5, containing an expression cassette with a CMV promoter, SARS-CoV-2 viral S protein gene, and adenosine monophosphate signal, 1*102 11 A virus particle.

[0363] 3. An immunobiological agent based on the genome of recombinant simian adenovirus serotype 25, containing an expression cassette with the CMV promoter, SARS-CoV-2 viral S protein gene, and adenosine monophosphate signal, 1*10 11 A virus particle.

[0364] Therefore, the stability of each adenovirus serotype included in the drug formulation was confirmed. The obtained drug formulations were stored at +2°C and +8°C for 3 months, then thawed; and changes in the titer of the recombinant adenovirus were assessed.

[0365] Table 2 - Composition of the experimental buffer solution

[0366] Table 2.

[0367]

[0368] The results of the experiments showed that the titer of recombinant adenovirus did not change after 3 months of storage in a buffer solution of lyophilized (freeze-dried) formulations for pharmaceutical preparations at +2°C and +8°C.

[0369] Therefore, the buffer solution for the lyophilized (freeze-dried) formulation developed for pharmaceutical preparations ensures the stability of all components of the developed pharmaceutical preparations within the following active range:

[0370] Tris: 0.0180% by mass to 0.0338% by mass;

[0371] Sodium chloride: 0.1044% by mass to 0.1957% by mass;

[0372] Sucrose: 5.4688% by mass to 10.2539% by mass;

[0373] Magnesium chloride hexahydrate: 0.0015% by mass to 0.0028% by mass;

[0374] EDTA: 0.0003% by mass to 0.0005% by mass;

[0375] Polysorbate-80: 0.0037% by mass to 0.0070% by mass;

[0376] Solvent: The remainder.

[0377] Example 8.

[0378] The effectiveness of immunization with the developed drug formulation is evaluated based on the assessment of humoral immune response.

[0379] One of the key characteristics of immunization effectiveness is antibody titer. This example brings up data related to changes in antibody titers against the SARS-CoV-2 glycoprotein on day 21 after administration of the drug formulation to experimental animals.

[0380] The experiment used female BALB / c mice, weighing 18g, a mammal species. All animals were divided into 31 groups of 5 animals each, with 10 animals per group. 8 Component 1 of the drug formulation was administered intramuscularly to animals at a dose of one virus particle / 100 μl, and two weeks later, 10 8 Component 2 was injected at a dose of 1 virus particle / 100 μl. Therefore, the following animal groups were formed:

[0381] 1) Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0382] 2) Ad26-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0383] 3) Ad26-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0384] 4) Ad26-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0385] 5) Ad26-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0386] 6) Ad26-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0387] 7) Ad26-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0388] 8) Ad26-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0389] 9) Ad26-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0390] 10) Ad26-null (component 1), Ad5-null (component 2);

[0391] 11) Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0392] 12) Ad26-CMV-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0393] 13) Ad26-CMV-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0394] 14) Ad26-CAG-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0395] 15) Ad26-CAG-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0396] 16) Ad26-CAG-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0397] 17) Ad26-EF1-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0398] 18) Ad26-EF1-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0399] 19) Ad26-EF1-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0400] 20) Ad26-null (component 1), simAd25-null (component 2);

[0401] 21) simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0402] 22) simAd25-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0403] 23) simAd25-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0404] 24) simAd25-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0405] 25) simAd25-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0406] 26) simAd25-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0407] 27) simAd25-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0408] 28) simAd25-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0409] 29) simAd25-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0410] 30) simAd25-null (component 1), Ad5-null (component 2);

[0411] 31) Phosphate buffered saline.

[0412] Three weeks later, blood samples were collected from the tail vein of the animals, and serum was separated. Antibody titers were measured using enzyme-linked immunosorbent assay (ELISA) according to the following protocol:

[0413] 1) At +4°C, the protein (S) was adsorbed into the wells of a 96-well ELISA plate for 16 hours.

[0414] 2) Then, to prevent nonspecific binding, the plate was “sealed” with 100 μl of 5% emulsion dissolved in TPBS per well. The plate was incubated in a shaker at 37°C for one hour.

[0415] 3) Serum samples from immunized mice were diluted using a 2-fold dilution method. A total of 12 dilutions were prepared for each sample.

[0416] 4) Add 50 μl of each diluted serum sample to the well of the plate.

[0417] 5) Then, incubate at 37°C for 1 hour.

[0418] 6) After incubation, wash the wells three times with phosphate buffer.

[0419] 7) Then, add a secondary antibody against mouse immunoglobulin conjugated with horseradish peroxidase.

[0420] 8) Next, incubate at 37°C for 1 hour.

[0421] 9) After incubation, wash the wells three times with phosphate buffer.

[0422] 10) Then, a tetramethylbenzidine (TMB) solution was added, which serves as the substrate for horseradish peroxidase and is converted into a colored compound via the reaction. The reaction was stopped after 15 minutes by adding sulfuric acid. Next, the optical density (OD) of the solution in each well was measured at a wavelength of 450 nm using a spectrophotometer.

[0423] The antibody titer was determined as the optical density of the solution was significantly higher than that of the negative control group at the last dilution. The results obtained (geometric mean) are presented in Table 3.

[0424] Table 3 - Antibody titers against S protein in mouse serum (geometric mean of antibody titers)

[0425] Table 3

[0426] 1 Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 33，779 2 Ad26-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2) 29，407 3 Ad26-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2) 33，779 4 Ad26-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 38，802 5 Ad26-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2) 38，802 6 Ad26-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2) 38，802 7 Ad26-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 33，779 8 Ad26-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2) 38，802 9 Ad26-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2) 33，779 10 Ad26-null (component 1), Ad5-null (component 2) 0 11 Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2) 38，802 12 Ad26-CMV-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2) 38，802 13 Ad26-CMV-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2) 33，779 14 Ad26-CAG-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2) 33，779 15 Ad26-CAG-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2) 33，779 16 Ad26-CAG-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2) 33，779 17 Ad26-EF1-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2) 38，802 18 Ad26-EF1-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2) 33，779 19 Ad26-EF1-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2) 33，779 20 Ad26-null (component 1), simAd25-null (component 2) 0 21 Ad25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 33，779 22 simAd25-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2) 29，407 23 simAd25-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2) 25，600 24 simAd25-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 33，779 25 simAd25-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2) 29，407 26 simAd25-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2) 29，407 27 simAd25-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 33，779 28 simAd25-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2) 38，802 29 simAd25-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2) 33，779 30 simAd25-null (component 1), Ad5-null (component 2) 0 31 Phosphate buffered saline 0

[0427] As shown in the presented data, all variants of the drug formulation induced a humoral immune response against the SARS-CoV-2 glycoprotein.

[0428] Example 9.

[0429] The efficacy of immunization with the developed drug formulation was evaluated compared to a control product containing a single serotype of recombinant adenovirus.

[0430] The purpose of this experiment was to compare the antibody titers against the SARS-CoV-2 virus S protein in the serum of mice after immunization with different variants of a developed drug formulation containing two different serotypes of recombinant adenovirus, with the antibody titers against the SARS-CoV-2 virus S protein in the serum of mice immunized twice with a control product containing one serotype of recombinant adenovirus.

[0431] In this experiment, 35 BalB / c mice, 18g each, were used.

[0432] Immunize the animals at 2-week intervals:

[0433] 1) Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2), 5*10 6 vp;

[0434] 2) Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2), 5*10 6 vp;

[0435] 3) simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2), 5*10 6 vp;

[0436] 4)Ad26-CMV-S-CoV2, Ad26-CMV-S-CoV2, 5*10 6 vp;

[0437] 5)Ad5-CMV-S-CoV2, Ad5-CMV-S-CoV2, 5*10 6 vp;

[0438] 6)simAd25-CMV-S-CoV2, simAd25-CMV-S-CoV2, 5*10 6 vp;

[0439] 7) PBS.

[0440] One month later, enzyme-linked immunosorbent assay (ELISA) was used to determine the antibody titer against the SARS-CoV-2 virus S antigen. The experimental results are presented in the table below.

[0441] Table 4 - Antibody titers against SARS-CoV-2 virus S antigen in mouse blood one month after immunization with the developed drug formulation and control product.

[0442] Table 4.

[0443] Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 3，104 Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2) 2，702 simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2) 3104 Ad26-CMV-S-CoV2, Ad26-CMV-S-CoV2 588 Ad5-CMV-S-CoV2, Ad5-CMV-S-CoV2 512 simAd25-CMV-S-CoV2, simAd25-CMV-S-CoV2 446 PBS 0

[0444] The data presented indicate that immunization of animals with the drug formulation enhances the immune response. This effect is demonstrated by significantly higher antibody titers in the serum of animals immunized with a drug formulation containing two carrier types, compared to the sum of antibody titers in groups immunized with a single carrier type.

[0445] Example 10.

[0446] The effectiveness of immunization with the developed drug formulation was evaluated based on the percentage of proliferating lymphocytes.

[0447] Cell-mediated immunity against SARS-CoV-2 virus was assessed by measuring the number of proliferating CD4+ and CD8+ lymphocytes in cultured mouse peripheral blood after a second restimulation of cells with a recombinant RBD fragment of the coronavirus S protein. To determine the number of proliferating CD4+ and CD8+ lymphocytes, a method using CFSE staining was employed. This method is based on the ability of fluorescent, non-toxic CFSE dyes to readily infiltrate cells. Upon stimulation with the antigen, lymphocytes begin to proliferate, and the dye from the parent cells is evenly distributed among the daughter cells. The labeling concentration, and therefore the fluorescence intensity in the daughter cells, is precisely reduced by two-fold. Therefore, dividing cells can be easily tracked by reducing fluorescence intensity.

[0448] C57BL / 6 mice were used in the experiment. All animals were divided into 31 groups (3 animals per group) and 10... 8 Component 1 of the drug formulation was administered intramuscularly at a dose of one viral particle / 100 μl. Two weeks later, at 10... 8 Component 2 was injected at a dose of 1 virus particle / 100 μl. Therefore, the following animal groups were formed:

[0449] 1) Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0450] 2) Ad26-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0451] 3) Ad26-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0452] 4) Ad26-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0453] 5) Ad26-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0454] 6) Ad26-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0455] 7) Ad26-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0456] 8) Ad26-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0457] 9) Ad26-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0458] 10) Ad26-null (component 1), Ad5-null (component 2);

[0459] 11) Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0460] 12) Ad26-CMV-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0461] 13) Ad26-CMV-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0462] 14) Ad26-CAG-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0463] 15) Ad26-CAG-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0464] 16) Ad26-CAG-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0465] 17) Ad26-EF1-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0466] 18) Ad26-EF1-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0467] 19) Ad26-EF1-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0468] 20) Ad26-null (component 1), simAd25-null (component 2);

[0469] 21) simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0470] 22) simAd25-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0471] 23) simAd25-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0472] 24) simAd25-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0473] 25) simAd25-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0474] 26) simAd25-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0475] 27) simAd25-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0476] 28) simAd25-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0477] 29) simAd25-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0478] 30) simAd25-null (component 1), Ad5-null (component 2);

[0479] 31) Phosphate buffered saline.

[0480] On day 8 of the experiment, the animals were euthanized. Lymphocytes were isolated from the spleen using a Ficoll-Urografin density gradient centrifugation method. Then, according to the technique (Monitoring lymphocyte proliferation in vitro and in vivo with the intracellular fluorescent dye carboxyfluorescein diacetate succinimidyl ester, BJ Quah et al., *Nature Protocols*, 2007, N...), lymphocyte proliferation was monitored in vitro and in vivo. o 2(9), 2049-2056) The isolated cells were stained with CFSE and cultured in the presence of antigen (SARS-CV-2 virus S glycoprotein).

[0481] The cells were then analyzed using a cellular fluorescence assay. The results were shown in... Figure 1 , 2 Therefore, it can be concluded that all variants of the developed drug formulation induce antigen-specific immune responses (CD4+ and CD8+).

[0482] Example 11.

[0483] The efficacy of the developed drug formulation against COVID-19 was evaluated in laboratory animals.

[0484] The protective efficacy of the developed drug formulation against COVID-19 was evaluated in Syrian golden hamsters with induced immunodeficiency using a model of lethal infection caused by the SARS-CoV-2 virus.

[0485] The animals were divided into 31 groups (8 animals per group) and immunized twice: using component 1 of the developed drug formulation (dose 10). 8 (1 virus particle / animal) and component 2 (dose 10) 8 (1 virus particle / animal), with an interval of 21 days.

[0486] Therefore, the following animal group was formed:

[0487] 1) Ad26-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0488] 2) Ad26-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0489] 3) Ad26-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0490] 4) Ad26-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0491] 5) Ad26-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0492] 6) Ad26-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0493] 7) Ad26-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0494] 8) Ad26-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0495] 9) Ad26-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0496] 10) Ad26-null (component 1), Ad5-null (component 2);

[0497] 11) Ad26-CMV-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0498] 12) Ad26-CMV-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0499] 13) Ad26-CMV-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0500] 14) Ad26-CAG-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0501] 15) Ad26-CAG-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2)

[0502] 16) Ad26-CAG-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0503] 17) Ad26-EF1-S-CoV2 (component 1), simAd25-CMV-S-CoV2 (component 2);

[0504] 18) Ad26-EF1-S-CoV2 (component 1), simAd25-CAG-S-CoV2 (component 2);

[0505] 19) Ad26-EF1-S-CoV2 (component 1), simAd25-EF1-S-CoV2 (component 2);

[0506] 20) Ad26-null (component 1), simAd25-null (component 2)

[0507] 21) simAd25-CMV-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0508] 22) simAd25-CMV-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0509] 23) simAd25-CMV-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0510] 24) simAd25-CAG-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0511] 25) simAd25-CAG-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0512] 26) simAd25-CAG-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0513] 27) simAd25-EF1-S-CoV2 (component 1), Ad5-CMV-S-CoV2 (component 2);

[0514] 28) simAd25-EF1-S-CoV2 (component 1), Ad5-CAG-S-CoV2 (component 2);

[0515] 29) simAd25-EF1-S-CoV2 (component 1), Ad5-EF1-S-CoV2 (component 2);

[0516] 30) simAd25-null (component 1), Ad5-null (component 2);

[0517] 31) Phosphate buffered saline.

[0518] Immunosuppressants were administered starting on day 7 after booster immunization; on day 14 after booster immunization, 10 mg / animal was given. 6 The TCID50 dose was determined by intranasal challenge with SARS-CoV-2 virus in animals at a dose of 50 μl.

[0519] During the experiment, the unvaccinated animals in the control group experienced a sharp decline in body weight after the challenge (on day 10, the average body weight decreased to 32% of baseline body weight). Meanwhile, during the first day after the challenge, the animals immunized with all variants of the drug formulation experienced a slight decrease in body weight, followed by an increase (on day 10, the average body weight increase was 11% of baseline body weight).

[0520] Figure 4The survival rate of the animals after the attack is shown. The results have demonstrated that immunization with all variants of the drug formulation provided 100% protection against lethal infection caused by the SARS-CoV-2 virus in animals with induced immunodeficiency. In the unvaccinated control group, the lethality rate was 100%.

[0521] Therefore, in a Syrian golden hamster model with induced immunodeficiency, it was demonstrated that the developed drug formulation induced a protective immune response, which ensured 100% protection of the animals from lethal infection caused by the SARS-CoV-2 virus.

[0522] Example 12:

[0523] Toxicity studies of the developed drug formulation

[0524] Toxicity was evaluated in sexually mature, distantly related male and female mice. Experimental studies were conducted using drug formulation variant 1 (component 1: Ad26-CMV-S-CoV2, component 2: Ad5-CMV-S-CoV2); drug formulation variant 2 (component 1: Ad26-CMV-S-CoV2, component 2: simAd25-CMV-S-CoV2); and drug formulation variant 3 (component 1: simAd25-CMV-S-CoV2, component 2: Ad5-CMV-S-CoV2). Each drug formulation component was administered intramuscularly and intravenously at escalating doses: 10 8 vp; 10 9 vp; 10 10 vp; and 10 11 vp.

[0525] No animal deaths or signs of poisoning were reported during the experiment. The vaccine was found to have no effect on the body weight or internal organ weight of the experimental animals. The structure of the internal organs in mice was not affected, as confirmed in a necropsy performed 14 days after administration of the vector-based vaccine. No local irritation was recorded in the experimental studies conducted.

[0526] Therefore, the research results indicate that the developed drug formulation is non-toxic.

[0527] Example 13.

[0528] Immunogenicity studies of the drug formulation developed based on variant 1 in primates

[0529] The purpose of this study was to assess the levels of humoral and T-cell-mediated immunity in primates following immunization with a developed drug formulation.

[0530] The evolution of humoral immune responses was assessed by elevating antibody titers against the SARS-CoV-2 virus S protein and virus-neutralizing antibody titers in primate blood. The evolution of cell-mediated immune responses was assessed by measuring the number of proliferating CD4+ and CD8+ T lymphocytes.

[0531] This study involved 17 male rhesus macaques, weighing between 2.0 kg and 2.2 kg. The animals were vaccinated with a developed drug formulation variant 1 (component 1: Ad26-CMV-S-CoV2, component 2: Ad5-CMV-S-CoV2). They were administered the human therapeutic dose of 10... 11 One viral particle / dose received component 1, and after 21 days, the human treatment dose was 10. 11 One virus particle / dose receiving component 2.

[0532] Blood samples were collected from all animals before vaccination (day 0) and on days 7, 14, and 28 post-vaccination. The titers of specific antibodies against the SARS-CoV-2 virus S protein RBD were measured in the serum of primates after immunization with the developed drug formulation using the following ELISA technique:

[0533] -Immobilize SARS-CoV-2 virus RBD antigen at a concentration of 100 ng / well on a plate;

[0534] - Perform a two-fold dilution of primate serum in blocking buffer (dilution ratio 1:50-1:51200) and incubate in wells of a plate (strip) with immobilized RBD antigen;

[0535] - After washing, the formed Ag-Ab complex was detected using a horseradish peroxidase-labeled conjugate specific to the Fc-fragment of the monkey antibody IgG (anti-monkey IgG (γ chain) (goat) antibody-617-101-012, ROCKLAND).

[0536] - After washing, a chromogenic substrate is added to the formed complex; then, a termination reagent is used to terminate the enzymatic reaction.

[0537] The color development (absorption) was recorded using a Multiskan FC (Thermo) spectrophotometer at two wavelengths: master filter -450nm and reference filter -620nm.

[0538] The titer of IgG antibodies against the SARS-CoV-2 virus S protein was defined as serum dilution, where, at the same dilution, the optical density value was twice higher than that of the negative control (serum from the same primate before drug administration). Experimental results were... Figure 5 As shown above.

[0539] These results indicate that antibody titers against SARS-CoV-2 increased in all animals immunized with the developed drug formulation. Peak antibody titers were then recorded one week after injection of component 2 (on day 28 of the experiment).

[0540] The levels of virus-neutralizing antibodies in the blood of rhesus monkeys were determined in a neutralization reaction based on the inhibition of SARS-CoV-2 virus-negative colonies in a one-day monolayer of Vero C1008 cells under agar-covered medium. The neutralization reaction was designed with a constant dose of virus-serum diluent.

[0541] The study included immune sera obtained from primates before vaccination (day 0) and on days 7, 14, and 28 after vaccination; positive control samples (serum samples from human convalescent patients containing specific antibodies against the SARS-CoV-2 virus); negative control samples (fetal bovine serum (FCS) containing no specific antibodies against the SARS-CoV-2 virus); and cultures of the SARS-CoV-2 virus.

[0542] Serum diluted 1:5 was used in the neutralization reaction. Working dilutions of SARS-CoV-2 virus-containing suspensions (“antigens”) were prepared in Hanks solution with 2% FCS and antibiotics (streptomycin sulfate and sodium benzylpenicillin), each at 100 U / ml, using serial decimal dilutions. The prepared dilutions achieved a SARS-CoV-2 virus concentration of 200 PFU / ml. -1 .

[0543] To conduct this experiment, a working surface area of ​​25 cm² was selected. 2 small plastic bottles For daily neutralization reactions with VeroC1008 cells, an equal volume mixture of serum and SARS-CoV-2 virus culture was incubated for 60 minutes between 36.5°C and 37.5°C, then added in 0.5 ml to a Vero C1008 cell monolayer (after initial removal of growth medium). After the antigen + antibody complex adhered to the cells for 60 minutes at 36.5–37.5°C, the inoculum was decanted. A primary agar cover designed for SARS-CoV-2 virus was then applied, and the monolayer was further incubated for 2 days between 36.5°C and 37.5°C.

[0544] Two days later, the infected cell monolayer was stained with a solution of 0.1% neutral red. For this purpose, a second agar cover layer was applied, and the cells were incubated at 36.5–37.5°C for 24 hours, and the number of negative colonies in the vials was counted. The antibody titer in the tested serum was defined as the highest dilution of serum in which the inhibition of the assay of SARS-CoV-2 virus-forming negative colonies was at least 50% higher than that in the negative control.

[0545] Studies have shown that on day 14 of the experiment, 17.6% of the animals had virus-neutralizing antibody levels higher than 1:5, while on day 28 of the experiment, 100% of the animals had virus-neutralizing antibody levels higher than that ratio.

[0546] Therefore, the results demonstrate that administration of the developed drug formulation induced a humoral immune response against the SARS-CoV-2 virus in primates.

[0547] To assess T-cell-mediated immune responses, monocytes were isolated from primate blood in a sucrose solution by density gradient centrifugation before vaccination (day 0) and on days 7, 14, and 28 after vaccination.

[0548] This method is based on the floating density gradient of blood cells. Peripheral blood cells can be separated and the monocyte fraction (MF) can be separated using density gradient centrifugation of a polysaccharide-sucrose solution in water. This fraction contains lymphocytes, subsets of monocytes, primitive hematopoietic cells, and fractions containing granulocytes and erythrocytes.

[0549] MF has a lower density than sucrose, so after centrifugation, it separates into layers on top of the sucrose reagent.

[0550] Granulocytes and erythrocytes are denser than the gradient density, and they cross the gradient and migrate to the bottom of the tube (Boyum A, Separation of leukocytes from blood and bonemarrow / / Scandinavian Journal of Clinical and Laboratory Investigations, 1968, Vol. 21, Supplement 97, pp. 1-9). When centrifuged at an appropriate speed, the smallest cells, platelets, remain in the serum and do not reach the "water / sucrose" phase interface.

[0551] After isolating mononuclear cells from peripheral blood of primates, the cells were stained with the fluorescent dye CFSE (Invivogen, USA) and placed in wells.

[0552] After seeding monocytes into wells, lymphocytes were restimulated in vitro by adding an RBD fragment of the coronavirus S protein (final protein concentration -1 μg / ml) to the culture medium. Intact cells without antigen were used as a negative control. The percentage of proliferating cells was measured 72 hours after antigen addition.

[0553] Experimental results show Figure 6 and 7 middle.

[0554] Experimental data showed that, on day 28 post-immunization, the maximum level of T-cell-mediated immunity induced by immunization with the developed drug formulation was recorded in primates, as assessed by the mean arithmetic value of the percentages of proliferating CD4+ T lymphocytes and CD8+ T lymphocytes. This result was correlated with a second (booster) immunization performed on day 21 of the study (1.2% vs. 0.1% in the unimmunized group). In this case, proliferating CD4+ and CD8+ T lymphocytes were re-stimulated for proliferation, increasing their percentage presence in the vaccinated animals.

[0555] In summary, it can be concluded that immunization of primates with the developed pharmaceutical formulation used in the tested doses and immunization regimens induced a significant (statistically significant difference from the values ​​in the control group of unimmunized animals) humoral immune response, characterized by an increase in antibody titers against the SARS-CoV-2 virus S protein and neutralizing antibody titers. It also induced T cell-mediated immunity involving CD4+ and CD8+ lymphocytes.

[0556] Example 14.

[0557] Cell-mediated immunity levels were assessed by measuring the number of proliferating CD4+ and CD8+ lymphocytes.

[0558] The immunogenicity of the developed drug formulation was evaluated by assessing the cell-mediated immune response to SARS-CoV-2 viral antigen in the blood of volunteers at different time points after vaccination.

[0559] Cell-mediated immune levels were evaluated in clinical trials of a drug formulation developed based on variant 1.

[0560] The trial involved 40 volunteers who received the following immunizations:

[0561] 1) Component 1 and 21 days later - use component 2 (component 1: Ad26-CMV-S-CoV2, component 2: Ad5-CMV-S-CoV2) of the liquid formulation variant 1 of the developed drug formulation, at a dose of 1x10 11 One virus particle (20 individuals).

[0562] 2) Component 1 and 21 days later - Component 2 (Component 1: Ad26-CMV-S-CoV2, Component 2: Ad5-CMV-S-CoV2) of the lyophilized (freeze-dried) formulation of the developed drug formulation variant 1, at a dose of 1x10 11 One virus particle (20 individuals).

[0563] Blood samples were collected from volunteers on day 0 (before vaccination), day 7, day 14, and day 28, and monocytes were isolated from the blood by density gradient centrifugation in a sucrose solution. The isolated cells were then stained with the fluorescent dye CFSE (Invivogen, USA) and seeded into wells.

[0564] Lymphocytes were then restimulated in vitro by adding coronavirus S protein (final protein concentration -1 μg / ml) to the culture medium. Intact cells without antigen were used as a negative control. The percentage of proliferating cells was determined 72 hours after antigen addition, and the culture medium was sampled for measuring gamma-interferon.

[0565] To determine the percentage of proliferating cells, antibodies were used to stain T lymphocytes for marker molecules CD3, CD4, and CD8 (anti-CD3 Pe-Cy7 (BD Biosciences, clone SK7), anti-CD4 APC (BD Biosciences, clone SK3), and anti-CD8 PerCP-Cy5.5 (BD Biosciences, clone SK1)). Proliferating (cells with lower levels of CFSE staining) CD4+ and CD8+ T lymphocytes in the cell mixture were measured using a high-performance cellular fluorometer BDFACS AriaIII (BD Biosciences, USA).

[0566] The percentage of proliferating cells obtained in each sample was determined by subtracting the results from the analysis of intact cells from the results obtained from the analysis of cells restimulated by the coronavirus S antigen. The results were displayed... Figure 8 and 9 (For liquid formulations of vaccines) and Figure 10 and 11 (Regarding freeze-dried formulations of vaccines).

[0567] According to the manufacturer's instructions, the concentration of interferon-gamma (IFNγ) in the culture medium of monocytes derived from human blood 72 hours after restimulation with coronavirus S protein was quantitatively determined using the "VECTOR-BEST" (Russia) kit. The received data showed... Figure 12 (For liquid formulations of vaccines) and Figure 13(Regarding freeze-dried formulations of vaccines).

[0568] The results of the study indicate that cell-mediated immunity induced by sequential immunization of volunteers with two formulations of drug formulation variant 1 (based on the median number of proliferating CD4+ and CD8+ T lymphocytes) increased with the number of days elapsed since the immunization date.

[0569] In both groups, peak values ​​of proliferating CD4+ and CD8+ T lymphocytes were recorded on day 28 post-immunization. The largest statistically significant difference in proliferating CD4+ and CD8+ T lymphocyte values ​​was reported between day 0 and day 28 of the study (p < 0.001).

[0570] based on Figure 12 and 13 The results shown lead to the conclusion that the level of cell-mediated immunity induced by sequential immunization of volunteers with two formulations of drug formulation variant 1 (based on median increase in IFNγ concentration) increases with the number of days elapsed since the immunization date.

[0571] The statistically significant difference in the increase in IFNγ concentration between pre-immunization (day 0) and day 14 post-immunization was p < 0.001. The maximum increase in IFNγ concentration was observed on day 28 post-immunization. The largest statistically significant difference (p < 0.001) in the increase in IFNγ concentration between day 0 and day 28 of the study was reported.

[0572] Therefore, based on these results, it can be concluded that immunization with the developed drug formulation can induce strong antigen-specific cell-mediated anti-infective immunity, as evidenced by high levels of statistical significance in parameters measured before and after immunization.

[0573] Example 15.

[0574] The immunogenicity of the developed drug formulation was assessed by evaluating antibody titers against SARS-CoV-2 viral antigens in the blood of volunteers at different time points after vaccination.

[0575] The trial involved 40 volunteers who received the following immunizations:

[0576] 1) Component 1 and 21 days later - use component 2 of the liquid formulation of the developed drug formulation variant 1 (component 1: Ad26-CMV-S-CoV2, component 2: Ad5-CMV-S-CoV2), at a dose of 1x10 11 One virus particle (20 individuals).

[0577] 2) Component 1 and 21 days later - Component 2 (Component 1: Ad26-CMV-S-CoV2, Component 2: Ad5-CMV-S-CoV2) of the lyophilized (freeze-dried) formulation of the developed drug formulation variant 1, at a dose of 1x10 11 One virus particle (20 individuals).

[0578] Blood samples were collected from volunteers on days 7, 14, and 28, and serum was separated from the blood.

[0579] Antibody titers against the SARS-CoV-2 viral S protein RBD were measured using an enzyme-linked immunosorbent assay (ELISA) and the "SARS-CoV-2-RBD-IFA-Gamaleya" test kit. The assay was performed according to the manufacturer's instructions.

[0580] exist Figure 14 The above shows the measured antibody titers against SARS-CoV-2 viral antigens in the serum of volunteers after receiving the liquid formulation of this product.

[0581] exist Figure 15 The above shows the measured results of antibody titers against SARS-CoV-2 virus antigens in the serum of volunteers after receiving the lyophilized (freeze-dried) formulation of this product.

[0582] As these results demonstrate, immunization of volunteers with the developed drug formulation (both liquid and lyophilized) facilitated strong humoral immunity (significantly different from the values ​​in the unimmunized control group), characterized by increased antibody titers against the SARS-CoV-2 viral S protein. Consequently, the level of the humoral immune response increased with the number of days elapsed from the immunization date.

[0583] Therefore, as demonstrated by the examples provided, the specified technical objectives have been achieved, particularly the development of agents that ensure effective induction of an immune response against the SARS-CoV-2 virus.

[0584] Industrial applicability

[0585] All the examples provided demonstrate the effectiveness and industrial applicability of pharmaceutical formulations that ensure effective induction of an immune response against the SARS-CoV-2 virus. sequence list <110> The Federal State Budgetary Institute of the Ministry of Health of the Russian Federation, named after Honorary Academician NF Gamaleya, is the National Research Center for Epidemiology and Microbiology. <120> Drug formulations used to induce specific immunity against SARS-CoV-2 <130> LHB2267520P <150> 2020127980 <151> 2020-08-22 <160> 7 <170> SIPOSequenceListing 1.0 <210> 1 <211> 4711 <212> DNA <213> Artificial Sequence <400> 1 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata 60 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat 120 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga 180 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc 240 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt 300 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat 360 gcggttttgg cagtacatca atggggcgtgg atagcggttt gactcacggg gatttccaag 420 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga 540 ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagagat ctggtaccgt cgacgcggcc gctcgagcct aagcttggta ccatgtttgt gttccttgtg ttattgccac 660 tagtctctag tcagtgtgtg aacctgacca caagaaccca gctgcctcca gcctacacca acagctttac cagaggcgtg tactaccccg acaaggtgtt cagatccagc gtgctgcact ctacccagga cctgttcctg cctttcttca gcaacgtgac ctggttccac gccatccacg 840 tgtccggcac caatggcacc aagagattcg acaacccgt gctgcccttc aacgacgggg tgtactttgc cagcaccgag aagtccaaca tcatcagagg ctggatcttc ggcaccacac tggacagcaa gacccagagc ctgctgatcg tgaacaacgc caccaacgtg gtcatcaaag tgtgcgagtt ccagttctgc aacgacccct tcctgggcgt ctactatcac aagaacaaca 1080 agagctggat ggaaagcgag ttccgggtgt acagcagcgc caacaactgc accttcgagt 1140 acgtgtccca gcctttcctg atggacctgg aaggcaagca gggcaacttc aagaacctgc 1200 gcgagttcgt gttcaagaac atcgacggct acttcaagat ctacagcaag cacaccccta 1260 tcaacctcgt gcgggatctg cctcagggct tctctgctct ggaacccctg gtggatctgc 1320 ccatcggcat caacatcacc cggtttcaga cactgctggc cctgcacaga agctacctga 1380 cacctggcga tagcagcagc ggatggacag ctggtgccgc cgcttactat gtgggctacc 1440 tgcagcctag aaccttcctg ctgaagtaca acgagaacgg caccatcacc gacgccgtgg 1500 attgtgctct ggatcctctg agcgagacaa agtgcaccct gaagtccttc accgtggaaa 1560 agggcatcta ccagaccagc aacttccggg tgcagcccac cgaatccatc gtgcggttcc 1620 ccaatatcac caatctgtgc cccttcggcg aggtgttcaa tgccaccaga ttcgcctctg 1680 tgtacgcctg gaaccggaag cggatcagca attgcgtggc cgactactcc gtgctgtaca 1740 actccgccag cttcagcacc ttcaagtgct acggcgtgtc ccctaccaag ctgaacgacc 1800 tgtgcttcac aaacgtgtac gccgacagct tcgtgatccg gggagatgaa gtgcggcaga 1860 ttgcccctgg acagacaggc aagatcgccg actacaacta caagctgccc gacgacttca 1920 ccggctgtgt gattgcctgg aacagcaaca acctggactc caaagtcggc ggcaactaca 1980 attacctgta ccggctgttc cggaagtcca atctgaagcc cttcgagcgg gacatctcca 2040 ccgagatcta tcaggccggc agcacccctt gtaacggcgt ggaaggcttc aactgctact 2100 tcccactgca gtcctacggc tttcagccca caaatggcgt gggctatcag ccctacagag 2160 tggtggtgct gagcttcgaa ctgctgcatg cccctgccac agtgtgcggc cctaagaaaa 2220 gcaccaatct cgtgaagaac aaatgcgtga acttcaactt caacggcctg accggcaccg 2280 gcgtgctgac agagagcaac aagaagttcc tgccattcca gcagtttggc cgggatattg 2340 ccgataccac agacgccgta cgagatcccc agacactgga aatcctggac atcacccctt 2400 gcagcttcgg cggagtgtct gtgatcaccc ctggcaccaa caccagcaat caggtggcag 2460 tgctgtacca ggacgtgaac tgtaccgaag tgcccgtggc cattcacgcc gatcagctga 2520 cacctacatg gcgggtgtac tccaccggca gcaatgtgtt tcagaccaga gccggctgtc 2580 tgatcggagc cgagcacgtg aacaatagct acgagtgcga catccccatc ggcgctggca 2640 tctgtgccag ctaccagaca cagacaaaca gccccagacg ggccagatct gtggccagcc 2700 agagcatcat tgcctacaca atgtctctgg gcgccgagaa cagcgtggcc tactccaaca 2760 actctatcgc tatccccacc aacttcacca tcagcgtgac cacagagatc ctgcctgtgt 2820 ccatgaccaa gaccagcgtg gactgcacca tgtacatctg cggcgattcc accgagtgct 2880 ccaacctgct gctgcagtac ggcagcttct gcacccagct gaatagagcc ctgacaggga 2940 tcgccgtgga acaggacaag aacacccaag aggtgttcgc ccaagtgaag cagatctaca 3000 agacccctcc tatcaaggac ttcggcggct tcaatttcag ccagattctg cccgatccta 3060 gcaagcccag caagcggagc ttcatcgagg acctgctgtt caacaaagtg acactggccg 3120 acgccggctt catcaagcag tatggcgatt gtctgggcga cattgccgcc agggatctga 3180 tttgcgccca gaagtttaac ggactgacag tgctgccacc actgctgacc gatgagatga 3240 tcgcccagta cacatctgcc ctgctggccg gcacaatcac aagcggctgg acatttggag 3300 ctggcgccgc tctgcagatc ccctttgcta tgcagatggc ctaccggttc aacggcatcg 3360 gagtgaccca gaatgtgctg tacgagaacc agaagctgat cgccaaccag ttcaacagcg 3420 ccatcggcaa gatccaggac agcctgagca gcacagcaag cgccctggga aagctgcagg 3480 acgtggtcaa ccagaatgcc caggcactga acaccctggt caagcagctg tcctccaact 3540 tcggcgccat cagctctgtg ctgaacgaca tcctgagcag actggacaag gtggaagccg 3600 aggtgcagat cgacagactg atcaccggaa ggctgcagtc cctgcagacc tacgttaccc 3660 agcagctgat cagagccgcc gagattagag cctctgccaa tctggccgcc accaagatgt 3720 ctgagtgtgt gctgggccag agcaagagag tggacttttg cggcaagggc taccacctga 3780 tgagcttccc tcagtctgcc cctcacggcg tggtgtttct gcacgtgaca tacgtgcccg 3840 ctcaagagaa gaatttcacc accgctccag ccatctgcca cgacggcaaa gcccactttc 3900 ctagagaagg cgtgttcgtg tccaacggca cccattggtt cgtgacccag cggaacttct 3960 acgagcccca gatcatcacc accgacaaca ccttcgtgtc tggcaactgc gacgtcgtga 4020 tcggcattgt gaacaatacc gtgtacgacc ctctgcagcc cgagctggac agcttcaaag 4080 agaactgga tagtacttt aagaaccaca caagccccga cgtggacctg ggcgacatca 4140 gcggaatcaa tgccagcgtc gtgaacatcc agaagagat cgaccggctg aacgaggtgg 4200 ccaagaatct gaacgagagc ctgatcgacc tgcaagaact ggggaagtac gagcagtaca 4260 tcaagtggcc ctggtacatc tggctggggct ttatcgccgg actgattgcc atcgtgatgg 4320 tcacaatcat gctgtgttgc atgaccagct gctgtagctg cctgaagggc tgttgtagct 4380 gtggcagctg ctgcaagttc gacgaggacg attctgagcc cgtgctcaaa ggagtcaaat 4440 tacattacac ataagatatc cgatccaccg gatctagata actgatcata atcagccata 4500 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga 4560 aaataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca 4620 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt 4680 gtggtttgtc caaactcatc aatgtatctt a 4711 <210> 2 <211> 5984 <212> DNA <213> Artificial Sequence <400> 2 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc 60 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca 120 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga 180 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc 240 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 300 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 360 tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact ctccccatct 420 ccccccctcc cacccccaat tttgtattta tttatttttt aattattttg tgcagcgatg 480 ggggcggggg gggggggcgc gcgccaggcg gggcggggcg gggcgagggg cggggcgggg 540 cgaggcggag aggtgcggcg gcagccaatc agagcggcgc gctccgaaag tttcctttta 600 tggcgaggcg gcggcggcgg cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg 660 ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc 720 tctgactgac cgcgttactc ccacaggtga gcgggcggga cggcccttct cctccgggct 780 gtaattagcg cttggtttaa tgacggcttg tttcttttct gtggctgcgt gaaagccttg 840 aggggctccg ggagggccct ttgtgcgggg ggagcggctc ggggggtgcg tgcgtgtgtg 900 tgtgcgtggg gagcgccgcg tgcggctccg cgctgcccgg cggctgtgag cgctgcgggc 960 gcggcgcggg gctttgtgcg ctccgcagtg tgcgcgaggg gagcgcggcc gggggcggtg 1020 ccccgcggtg cgggggggct gcgaggggaa caaaggctgc gtgcggggtg tgtgcgtggg 1080 gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca cccccctccc 1140 cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg 1200 ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct 1260 cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga 1320 ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt 1380 cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc 1440 gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt 1500 gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac 1560 ggctgccttc ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg accggcggct 1620 ctagaaagct tggtaccatg tttgtgttcc ttgtgttatt gccactagtc tctagtcagt 1680 gtgtgaacct gaccacaaga acccagctgc ctccagccta caccaacagc tttaccagag 1740 gcgtgtacta ccccgacaag gtgttcagat ccagcgtgct gcactctacc caggacctgt 1800 tcctgccttt cttcagcaac gtgacctggt tccacgccat ccacgtgtcc ggcaccaatg 1860 gcaccaagag attcgacaac cccgtgctgc ccttcaacga cggggtgtac tttgccagca 1920 ccgagaagtc caacatcatc agaggctgga tcttcggcac cacactggac agcaagaccc 1980 agagcctgct gatcgtgaac aacgccacca acgtggtcat caaagtgtgc gagttccagt 2040 tctgcaacga ccccttcctg ggcgtctact atcacaagaa caacaagagc tggatggaaa 2100 gcgagttccg ggtgtacagc agcgccaaca actgcacctt cgagtacgtg tcccagcctt 2160 tcctgatgga cctggaaggc aagcagggca acttcaagaa cctgcgcgag ttcgtgttca 2220 agaacatcga cggctacttc aagatctaca gcaagcacac ccctatcaac ctcgtgcggg 2280 atctgcctca gggcttctct gctctggaac ccctggtgga tctgcccatc ggcatcaaca 2340 tcacccggtt tcagacactg ctggccctgc acagaagcta cctgacacct ggcgatagca 2400 gcagcggatg gacagctggt gccgccgctt actatgtggg ctacctgcag cctagaacct 2460 tcctgctgaa gtacaacgag aacggcacca tcaccgacgc cgtggattgt gctctggatc 2520 ctctgagcga gacaaagtgc accctgaagt ccttcaccgt ggaaaagggc atctaccaga 2580 ccagcaactt ccgggtgcag cccaccgaat ccatcgtgcg gttccccaat atcaccaatc 2640 tgtgcccctt cggcgaggtg ttcaatgcca ccagattcgc ctctgtgtac gcctggaacc 2700 ggaagcggat cagcaattgc gtggccgact actccgtgct gtacaactcc gccagcttca 2760 gcaccttcaa gtgctacggc gtgtccccta ccaagctgaa cgacctgtgc ttcacaaacg 2820 tgtacgccga cagcttcgtg atccggggag atgaagtgcg gcagattgcc cctggacaga 2880 caggcaagat cgccgactac aactacaagc tgcccgacga cttcaccggc tgtgtgattg 2940 cctggaacag caacaacctg gactccaaag tcggcggcaa ctacaattac ctgtaccggc 3000 tgttccggaa gtccaatctg aagcccttcg agcgggacat ctccaccgag atctatcagg 3060 ccggcagcac cccttgtaac ggcgtggaag gcttcaactg ctacttccca ctgcagtcct 3120 acggctttca gcccacaaat ggcgtgggct atcagcccta cagagtggtg gtgctgagct 3180 tcgaactgct gcatgcccct gccacagtgt gcggccctaa gaaaagcacc aatctcgtga 3240 agaacaaatg cgtgaacttc aacttcaacg gcctgaccgg caccggcgtg ctgacagaga 3300 gcaacaagaa gttcctgcca ttccagcagt ttggccggga tattgccgat accacagacg 3360 ccgtacgaga tccccagaca ctggaaatcc tggacatcac cccttgcagc ttcggcggag 3420 tgtctgtgat cacccctggc accaacacca gcaatcaggt ggcagtgctg taccaggacg 3480 tgaactgtac cgaagtgccc gtggccattc acgccgatca gctgacacct acatggcggg 3540 tgtactccac cggcagcaat gtgtttcaga ccagagccgg ctgtctgatc ggagccgagc 3600 acgtgaacaa tagctacgag tgcgacatcc ccatcggcgc tggcatctgt gccagctacc 3660 agacacagac aaacagcccc agacgggcca gatctgtggc cagccagagc atcattgcct 3720 acacaatgtc tctgggcgcc gagaacagcg tggcctactc caacaactct atcgctatcc 3780 ccaccaactt caccatcagc gtgaccacag agatcctgcc tgtgtccatg accaagacca 3840 gcgtggactg caccatgtac atctgcggcg attccaccga gtgctccaac ctgctgctgc 3900 agtacggcag cttctgcacc cagctgaata gagccctgac agggatcgcc gtggaacagg 3960 acaagaacac ccaagaggtg ttcgcccaag tgaagcagat ctacaagacc cctcctatca 4020 aggacttcgg cggcttcaat ttcagccaga ttctgcccga tcctagcaag cccagcaagc 4080 ggagcttcat cgaggacctg ctgttcaaca aagtgacact ggccgacgcc ggcttcatca 4140 agcagtatgg cgattgtctg ggcgacattg ccgccaggga tctgatttgc gcccagaagt 4200. ttaacggact gacagtgctg ccaccactgc tgaccgatga gatgatcgcc cagtacacat 4320. ctgccctgct ggccggcaca atcacaagcg gctggacatt tggagctggc gccgctctgc agatcccctt tgctatgcag atggcctacc ggttcaacgg catcggagtg acccagaatg 4440. tgctgtacga gaaccagaag ctgatcgcca accagttcaa cagcgccatc ggcaagatcc aggacagcct gagcagcaca gcaagcgccc tgggaaagct gcaggacgtg gtcaaccaga atgcccaggc actgaacacc ctggtcaagc agctgtcctc caacttcggc gccatcagct 4560 ctgtgctgaa cgacatcctg agcagactgg acaaggtgga agccgaggtg cagatcgaca gactgatcac cggaaggctg cagtccctgc agacctacgt tacccagcag ctgatcagag ccgccgagat tagagcctct gccaatctgg ccgccaccaa gatgtctgag tgtgtgctgg 4740 gccagagcaa gagagtggac ttttgcggca agggctacca cctgatgagc ttccctcagt ctgcccctca cggcgtggtg tttctgcacg tgacatacgt gcccgctcaa gagaagaatt tcaccaccgc tccagccatc tgccacgacg gcaaagccca ctttcctaga gaaggcgtgt 4920 tcgtgtccaa cggcacccat tggttcgtga cccagcggaa cttctacgag ccccagatca 4980 tcaccaccga caacaccttc gtgtctggca actgcgacgt cgtgatcggc attgtgaaca 5040 ataccgtgta cgaccctctg cagcccgagc tggacagctt caaagaggaa ctggataagt 5100 actttaagaa ccacacaagc cccgacgtgg acctgggcga catcagcgga atcaatgcca 5160 gcgtcgtgaa catccagaaa gagatcgacc ggctgaacga ggtggccaag aatctgaacg 5220 agagcctgat cgacctgcaa gaactgggga agtacgagca gtacatcaag tggccctggt 5280 acatctggct gggctttatc gccggactga ttgccatcgt gatggtcaca atcatgctgt 5340 gttgcatgac cagctgctgt agctgcctga agggctgttg tagctgtggc agctgctgca 5400 agttcgacga ggacgattct gagcccgtgc tcaaaggagt caaattacat tacacataat 5460 tcactcctca ggtgcaggct gcctatcaga aggtggtggc tggtgtggcc aatgccctgg 5520 ctcacaaata ccactgagat ctttttccct ctgccaaaaa ttatggggac atcatgaagc 5580 cccttgagca tctgacttct ggctaataaa ggaaatttat tttcattgca atagtgtgtt 5640 ggaatttttt gtgtctctca ctcggaagga catatgggag ggcaaatcat ttaaaacatc 5700 agaatgagta tttggtttag agtttggcaa catatgccca tatgctggct gccatgaaca 5760 aaggttggct ataaagaggt catcagtata tgaaacagcc ccctgctgtc cattccttat 5820 tccatagaaa agccttgact tgaggttaga tttttttata ttttgttttg tgttattttt 5880 tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt cctcctctcc 5940 tgactactcc cagtcatagc tgtccctctt ctcttatgga gatc 5984 <210> 3 <211> 5314 <212> DNA <213> Artificial Sequence <400> 3 ggtgaggctc cggtgcccgt cagtgggcag agcgcacatc gcccacagtc cccgagaagt 60 tggggggagg ggtcggcaat tgaaccggtg cctagagaag gtggcgcggg gtaaactggg 120 aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa ccgtatataa 180 gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt tgccgccaga acacaggtaa 240 gtgccgtgtg tggttcccgc gggcctggcc tctttacggg ttatggccct tgcgtgcctt 300 gaattacttc cacctggctg cagtacgtga ttcttgatcc cgagcttcgg gttggaagtg 360 ggtgggagag ttcgaggcct tgcgcttaag gagccccttc gcctcgtgct tgagttgagg 420 cctggcctgg gcgctggggc cgccgcgtgc gaatctggtg gcaccttcgc gcctgtctcg 480 ctgctttcga taagtctcta gccatttaaa atttttgatg acctgctgcg acgctttttt 540 tctggcaaga tagtcttgta aatgcgggcc aagatctgca cactggtatt tcggtttttg 600 gggccgcggg cggcgacggg gcccgtgcgt cccagcgcac atgttcggcg aggcggggcc 660 tgcgagcgcg gccaccgaga atcggacggg ggtagtctca agctggccgg cctgctctgg 720 tgcctggcct cgcgccgccg tgtatcgccc cgccctgggc ggcaaggctg gcccggtcgg 780 caccagttgc gtgagcggaa agatggccgc ttcccggccc tgctgcaggg agctcaaaat 840 ggaggacgcg gcgctcggga gagcgggcgg gtgagtcacc cacacaaagg aaaagggcct 900 ttccgtcctc agccgtcgct tcatgtgact ccacggagta ccgggcgccg tccaggcacc 960 tcgattagtt ctcgagcttt tggagtacgt cgtctttagg ttggggggag gggttttatg 1020 cgatggagtt tccccacact gagtgggtgg agactgaagt taggccagct tggcacttga 1080 tgtaattctc cttggaattt gccctttttg agtttggatc ttggttcatt ctcaagcctc 1140 agacagtggt tcaaagtttt tttcttccat ttcaggtgtc gtgaggaatt agcttggtac 1200 taatacgact cacaagcttg gtaccatgtt tgtgttcctt gtgttattgc cactagtctc 1260 tagtcagtgt gtgaacctga ccacaagaac ccagctgcct ccagcctaca ccaacagctt 1320 taccagaggc gtgtactacc ccgacaaggt gttcagatcc agcgtgctgc actctaccca 1380 ggacctgttc ctgcctttct tcagcaacgt gacctggttc cacgccatcc acgtgtccgg 1440 caccaatggc accaagagat tcgacaaccc cgtgctgccc ttcaacgacg gggtgtactt 1500 tgccagcacc gagaagtcca acatcatcag aggctggatc ttcggcacca cactggacag 1560 caagacccag agcctgctga tcgtgaacaa cgccaccaac gtggtcatca aagtgtgcga 1620 gttccagttc tgcaacgacc ccttcctggg cgtctactat cacaagaaca acaagagctg 1680 gatggaaagc gagttccggg tgtacagcag cgccaacaac tgcaccttcg agtacgtgtc 1740 ccagcctttc ctgatggacc tggaaggcaa gcagggcaac ttcaagaacc tgcgcgagtt 1800 cgtgttcaag aacatcgacg gctacttcaa gatctacagc aagcacaccc ctatcaacct 1860 cgtgcgggat ctgcctcagg gcttctctgc tctggaaccc ctggtggatc tgcccatcgg 1920 catcaacatc acccggtttc agacactgct ggccctgcac agaagctacc tgacacctgg 1980 cgatagcagc agcggatgga cagctggtgc cgccgcttac tatgtgggct acctgcagcc 2040 tagaaccttc ctgctgaagt acaacgagaa cggcaccatc accgacgccg tggattgtgc 2100 tctggatcct ctgagcgaga caaagtgcac cctgaagtcc ttcaccgtgg aaaagggcat 2160 ctaccagacc agcaacttcc gggtgcagcc caccgaatcc atcgtgcggt tccccaatat 2220 caccaatctg tgccccttcg gcgaggtgtt caatgccacc agattcgcct ctgtgtacgc 2280 ctggaaccgg aagcggatca gcaattgcgt ggccgactac tccgtgctgt acaactccgc 2340 cagcttcagc accttcaagt gctacggcgt gtcccctacc aagctgaacg acctgtgctt 2400 cacaaacgtg tacgccgaca gcttcgtgat ccggggagat gaagtgcggc agattgcccc 2460 tggacagaca ggcaagatcg ccgactacaa ctacaagctg cccgacgact tcaccggctg 2520 tgtgattgcc tggaacagca acaacctgga ctccaaagtc ggcggcaact acaattacct 2580 gtaccggctg ttccggaagt ccaatctgaa gcccttcgag cgggacatct ccaccgagat 2640 ctatcaggcc ggcagcaccc cttgtaacgg cgtggaaggc ttcaactgct acttcccact 2700 gcagtcctac ggctttcagc ccacaaatgg cgtgggctat cagccctaca gagtggtggt 2760 gctgagcttc gaactgctgc atgcccctgc cacagtgtgc ggccctaaga aaagcaccaa 2820 tctcgtgaag aacaaatgcg tgaacttcaa cttcaacggc ctgaccggca ccggcgtgct 2880 gacagagagc aacaagaagt tcctgccatt ccagcagttt ggccgggata ttgccgatac 2940 cacagacgcc gtacgagatc cccagacact ggaaatcctg gacatcaccc cttgcagctt 3000 cggcggagtg tctgtgatca cccctggcac caacaccagc aatcaggtgg cagtgctgta 3060 ccaggacgtg aactgtaccg aagtgcccgt ggccattcac gccgatcagc tgacacctac 3120 atggcgggtg tactccaccg gcagcaatgt gtttcagacc agagccggct gtctgatcgg 3180 agccgagcac gtgaacaata gctacgagtg cgacatcccc atcggcgctg gcatctgtgc 3240 cagctaccag acacagacaa acagccccag acgggccaga tctgtggcca gccagagcat 3300 cattgcctac acaatgtctc tgggcgccga gaacagcgtg gcctactcca acaactctat 3360 cgctatcccc accaacttca ccatcagcgt gaccacagag atcctgcctg tgtccatgac 3420 caagaccagc gtggactgca ccatgtacat ctgcggcgat tccaccgagt gctccaacct 3480 gctgctgcag tacggcagct tctgcaccca gctgaataga gccctgacag ggatcgccgt 3540 ggaacaggac aagaacaccc aagaggtgtt cgcccaagtg aagcagatct acaagacccc 3600 tcctatcaag gacttcggcg gcttcaattt cagccagatt ctgcccgatc ctagcaagcc 3660 cagcaagcgg agcttcatcg aggacctgct gttcaacaaa gtgacactgg ccgacgccgg 3720 cttcatcaag foottggcg attgtctggg cgacattgcc gccagggatc tgatttgcgc 3780 ccagaagttt aacggactga cagtgctgcc accactgctg accgatgaga tgatcgccca 3840 gtacacatct gccctgctgg ccggcacaat caaagcggc tggacatttg gagctggcgc 3900 cgctctgcag atcccctttg ctatgcagat ggcctaccgg ttcaacggca tcggagtgac 3960 ccagaatgtg ctgtacgaga accagaagct gatcgccaac cagttcaaca gcgccatcgg 4020 caagatccag gacagcctga gcagcacagc aagcgccctg ggaaagctgc aggacgtggt 4080 caaccagaat gcccaggcac tgaacaccct ggtcaagcag ctgtcctcca acttcggcgc 4140 catcagctct gtgctgaacg acatcctgag cagactggac aaggtggaag ccgaggtgca 4200 gatcgacaga ctgatcaccg gaaggctgca gtccctgcag acctacgtta cccagcagct 4260 gatcagagcc gccgagatta gagcctctgc caatctggcc gccaccaaga tgtctgagtg 4320 tgtgctgggc cagagcaaga gagtggactt ttgcggcaag ggctaccacc tgatgagctt 4380 ccctcagtct gcccctcacg gcgtggtgtt tctgcacgtg acatacgtgc ccgctcaaga 4440 gaagaatttc accaccgctc cagccatctg ccacgacgg aaagcccact ttcctagaga 4500 aggcgtgttc gtgtccaacg gcacccattg gttcgtgacc cagcggaact tctacgagcc 4560 ccagatcatc accaccgaca acaccttcgt gtctggcaac tgcgacgtcg tgatcggcat 4620 tgtgaacaat accgtgtacg accctctgca gcccgagctg gacagcttca aaggaact 4680 ggataagtac tttaagaacc acacaagccc cgacgtggac ctgggcgaca tcagcggaat 4740 caatgccagc gtcgtgaaca tccagaaaga gatcgaccgg ctgaacgagg tggccaagaa 4800 tctgaacgag agcctgatcg acctgcaaga actgggggaag tacgagcagt acatcaagtg 4860 gccctggtac atctggctgg gctttatcgc cggactgatt gccatcgtga tggtcacaat 4920 catgctgtgt tgcatgacca gctgctgtag ctgcctgaag ggctgttgta gctgtggcag 4980 ctgctgcaag ttcgacgagg acgattctga gcccgtgctc aaaggagtca aattacatta 5040 cakataagat ctagagtcgg ggcggccggc cgctcgctga tcagcctcga ctgtgccttc 5100 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc 5160 cactccact gtcctttcct ataaatga ggaattgca tcgcattgtc tgagtaggtg 5220 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa 5280 tagcaggcat gctggggatc cgagtgtcga taag 5314 <210> 4 <211> 4678 <212> DNA <213> Artificial Sequence <400> 4 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata 60 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat 120 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga 180 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc 240 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt 300 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat 360 gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag 420 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc 480 aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga 540 ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagagat ctggtaccat 600 gtttgtgttc cttgtgttat tgccactagt ctctagtcag tgtgtgaacc tgaccacaag 660 aacccagctg cctccagcct acaccaacag ctttaccaga ggcgtgtact accccgacaa 720 ggtgttcaga tccagcgtgc tgcactctac ccaggacctg ttcctgcctt tcttcagcaa 780 cgtgacctgg ttccacgcca tccacgtgtc cggcaccaat ggcaccaaga gattcgacaa 840 ccccgtgctg cccttcaacg acggggtgta ctttgccagc accgagaagt ccaacatcat 900 cagaggctgg atcttcggca ccacactgga cagcaagacc cagagcctgc tgatcgtgaa 960 caacgccacc aacgtggtca tcaaagtgtg cgagttccag ttctgcaacg accccttcct 1020 gggcgtctac tatcacaaga acaacaagag ctggatggaa agcgagttcc gggtgtacag 1080 cagcgccaac aactgcacct tcgagtacgt gtcccagcct ttcctgatgg acctggaagg 1140 caagcagggc aacttcaaga acctgcgcga gttcgtgttc aagaacatcg acggctactt 1200 caagatctac agcaagcaca cccctatcaa cctcgtgcgg gatctgcctc agggcttctc 1260 tgctctggaa cccctggtgg atctgcccat cggcatcaac atcacccggt ttcagacact 1320 gctggccctg cacagaagct acctgacacc tggcgatagc agcagcggat ggacagctgg 1380 tgccgccgct tactatgtgg gctacctgca gcctagaacc ttcctgctga agtacaacga 1440 gaacggcacc atcaccgacg ccgtggattg tgctctggat cctctgagcg agacaaagtg 1500 caccctgaag tccttcaccg tggaaaaggg catctaccag accagcaact tccgggtgca 1560 gcccaccgaa tccatcgtgc ggttccccaa tatcaccaat ctgtgcccct tcggcgaggt 1620 gttcaatgcc accagattcg cctctgtgta cgcctggaac cggaagcgga tcagcaattg 1680 cgtggccgac tactccgtgc tgtacaactc cgccagcttc agcaccttca agtgctacgg 1740 cgtgtcccct accaagctga acgacctgtg cttcacaaac gtgtacgccg acagcttcgt 1800 gatccgggga gatgaagtgc ggcagattgc ccctggacag acaggcaaga tcgccgacta 1860 caactacaag ctgcccgacg acttcaccgg ctgtgtgatt gcctggaaca gcaacaacct 1920 ggactccaaa gtcggcggca actacaatta cctgtaccgg ctgttccgga agtccaatct 1980 gaagcccttc gagcgggaca tctccaccga gatctatcag gccggcagca ccccttgtaa 2040 cggcgtggaa ggcttcaact gctacttccc actgcagtcc tacggctttc agcccacaaa 2100 tggcgtgggc tatcagccct acagagtggt ggtgctgagc ttcgaactgc tgcatgcccc 2160 tgccacagtg tgcggcccta agaaaagcac caatctcgtg aagaacaaat gcgtgaactt 2220 caacttcaac ggcctgaccg gcaccggcgt gctgacagag agcaacaaga agttcctgcc 2280 attccagcag tttggccggg atattgccga taccacagac gccgtacgag atccccagac 2340 actggaaatc ctggacatca ccccttgcag cttcggcgga gtgtctgtga tcacccctgg 2400 caccaacacc agcaatcagg tggcagtgct gtaccaggac gtgaactgta ccgaagtgcc 2460 cgtggccatt cacgccgatc agctgacacc tacatggcgg gtgtactcca ccggcagcaa 2520 tgtgtttcag accagagccg gctgtctgat cggagccgag cacgtgaaca atagctacga 2580 gtgcgacatc cccatcggcg ctggcatctg tgccagctac cagacacaga caaacagccc 2640 cagacgggcc agatctgtgg ccagccagag catcattgcc tacacaatgt ctctgggcgc 2700 cgagaacagc gtggcctact ccaacaactc tatcgctatc cccaccaact tcaccatcag 2760 cgtgaccaca gagatcctgc ctgtgtccat gaccaagacc agcgtggact gcaccatgta 2820 catctgcggc gattccaccg agtgctccaa cctgctgctg cagtacggca gcttctgcac 2880 ccagctgaat agagccctga cagggatcgc cgtggaacag gacaagaaca cccaagaggt 2940 gttcgcccaa gtgaagcaga tctacaagac ccctcctatc aaggacttcg gcggcttcaa 3000 tttcagccag attctgcccg atcctagcaa gcccagcaag cggagcttca tcgaggacct 3060 gctgttcaac aaagtgacac tggccgacgc cggcttcatc aagcagtatg gcgattgtct 3120 gggcgacatt gccgccaggg atctgatttg cgcccagaag tttaacggac tgacagtgct 3180 gccaccactg ctgaccgatg agatgatcgc ccagtacaca tctgccctgc tggccggcac 3240 aatcacaagc ggctggacat ttggagctgg cgccgctctg cagatcccct ttgctatgca 3300 gatggcctac cggttcaacg gcatcggagt gacccagaat gtgctgtacg agaaccagaa 3360 gctgatcgcc aaccagttca acagcgccat cggcaagatc caggacagcc tgagcagcac 3420 agcaagcgcc ctgggaaagc tgcaggacgt ggtcaaccag aatgcccagg cactgaacac 3480 cctggtcaag cagctgtcct ccaacttcgg cgccatcagc tctgtgctga acgacatcct 3540 gaccagactg gacaaggtgg aagccgaggt gcagatcgac agactgatca ccggaaggct 3600 gcagtccctg cagacctacg ttacccagca gctgatcaga gccgccgaga ttagagcctc 3660 3720 cttttgcggc aagggctacc acctgatgag cttccctcag tctgcccctc acggcgtggt 3780 gtttctgcac gtgacatacg tgcccgctca agagaagaat ttcaccaccg ctccagccat 3840 ctgccacgac ggcaaagccc actttcctag agaaggcgtg ttcgtgtcca acggcaccca 3900 ttggttcgtg acccagcgga acttctacga gccccagatc atcaccaccg acaaccctt 3960 cgtgtctggc aactgcgacg tcgtgatcgg cattgtgaac aataccgtgt acgaccctct 4020 gcagcccgag ctggacagct tcaaaagaga actggataag tactttaaga accacacaag 4080 ccccgacgtg gacctgggcg acatcagcgg aatcaatgcc agcgtcgtga acatccagaa 4140 agagatcgac cggctgaacg aggtggccaa gaatctgaac gagagcctga tcgacctgca 4200 agaactgggg aagtacgagc agtacatcaa gtggccctgg tacatctggc tgggcttat 4260 cgccggactg attgccatcg tgatggtcac aatcatgctg tgttgcatga ccagctgctg 4320 tagctgcctg aagggctgtt gtagctgtgg cagctgctgc aagttcgacg aggacgattc 4380 tgagcccgtg ctcaaaggag tcaaattaca ttacacataa gatatcgcgg ccgctcgagt 4440 ctagataact gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa 4500 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact 4560 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata 4620 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatctta 4678 <210> 5 <211> 33765 <212> DNA <213> Recombinant human adenovirus serotype 26 <400> 5 catcatcaat aatacccc acaaagtaaa caaaagttaa tatgcaaatg agcttttgaa 60 ttttaacggt tttggggcgg agccaacgct gattggacga gaaacggtga tgcaaatgac 120 gtcacgacgc acggctaacg gtcgccgcgg aggcgtggcc tagcccggaa gcaagtcgcg 180 gggctgatga cgtataaaaa agcggacttt agacccggaa acggccgatt ttcccgcggc 240 cacgcccgga tatgaggtaa ttctgggcgg atgcaagtga aattaggtca ttttggcgcg 300 aaaactgaat gaggaagtga aaagcgaaaa ataccggtcc ctcccagggc ggaatattta 360 ccgagggccg agagactttg accgattacg tgggggtttc gattgcggtg tttttttcgc 420 gaatttccgc gtccgtgtca aagtccggtg tttatgtcac agatcagctg gtttccttta 480 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt 540 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt 600 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt 660 taaagcaagt aaaacctcta caaatgtggt atggctgatt atgatcagtt atctagatcc 720 ggtggatcgg atatcttatg tgtaatgtaa tttgactcct ttgagcacgg gctcagaatc 780 gtcctcgtcg aacttgcagc agctgccaca gctacaacag cccttcaggc agctacagca 840 gctggtcatg caacacagca tgattgtgac catcacgatg gcaatcagtc cggcgataaa 900 gcccagccag atgtaccagg gccacttgat gtactgctcg tacttcccca gttcttgcag 960 gtcgatcagg ctctcgttca gattcttggc cacctcgttc agccggtcga tctctttctg 1020 gatgttcacg acgctggcat tgattccgct gatgtcgccc aggtccacgt cggggcttgt 1080 gtggttctta aagtacttat ccagttcctc tttgaagctg tccagctcgg gctgcagagg 1140 gtcgtacacg gtattgttca caatgccgat cacgacgtcg cagttgccag acacgaaggt 1200 gttgtcggtg gtgatgatct ggggctcgta gaagttccgc tgggtcacga accaatgggt 1260 gccgttggac acgaacacgc cttctctagg aaagtgggct ttgccgtcgt ggcagatggc 1320 tggagcggtg gtgaaattct tctcttgagc gggcacgtat gtcacgtgca gaaacaccac 1380 gccgtgaggg gcagactgag ggaagctcat caggtggtag cccttgccgc aaaagtccac 1440 tctcttgctc tggcccagca cacactcaga catcttggtg gcggccagat tggcagaggc 1500 tctaatctcg gcggctctga tcagctgctg ggtaacgtag gtctgcaggg actgcagcct tccggtgatc agtctgtcga tctgcacctc ggcttccacc ttgtccagtc tgctcaggat 1620 gtcgttcagc acagagctga tggcgccga gttggaggac agctgcttga ccagggtgtt 1680. gtcgttcagc cagtgcctgg gcattctggt tgaccacgtc ctgcagcttt cccagggcgc ttgctgtgct 1740 gctcaggctg tcctggatct tgccgatggc gctgttgaac tggttggcga tcagcttctg gttctcgtac agcacattct gggtcactcc gatgccgttg aaccggtagg ccatctgcat agcaaagggg atctgcagag cggcgccagc tccaaatgtc cagccgcttg tgattgtgcc ggccagcagg gcagatgtgt actgggcgat catctcatcg gtcagcagtg gtggcagcac tgtcagtccg ttaaacttct gggcgcaaat cagatccctg gcggcaatgt cgcccagaca atcgccatac tgcttgatga agccggcgtc ggccagtgtc actttgttga acagcaggtc ctcgatgaag ctccgcttgc tgggcttgct aggatcgggc agaatctggc tgaaattgaa gccgccgaag tccttgatag gaggggtctt gtagatctgc ttcacttggg cgaacacctc ttgggtgttc ttgtcctgtt ccacggcgat ccctgtcagg gctctattca gctgggtgca 2280 gaagctgccg tactgcagca gcaggttgga gcactcggtg gaatcgccgc agatgtacat 2340 ggtgcagtcc acgctggtct tggtcatgga cacaggcagg atctctgtgg tcacgctgat 2400 ggtgaagttg gtggggatag cgatagagtt gttggagtag gccacgctgt tctcggcgcc 2460 cagagacatt gtgtaggcaa tgatgctctg gctggccaca gatctggccc gtctggggct 2520 gtttgtctgt gtctggtagc tggcacagat gccagcgccg atggggatgt cgcactcgta 2580 gctattgttc acgtgctcgg ctccgatcag acagccggct ctggtctgaa acacattgct 2640 gccggtggag tacacccgcc atgtaggtgt cagctgatcg gcgtgaatgg ccacgggac 2700 ttcggtacag ttcacgtcct ggtacagcac tgccacctga ttgctggtgt tggtgccagg 2760 ggtgatcaca gacactccgc cgaagctgca aggggtgatg tccaggattt ccagtgtctg 2820 gggatctcgt acggcgtctg tggtatcggc aatatcccgg ccaaactgct ggaatggcag 2880 gaacttcttg ttgctctctg tcagcacgcc ggtgccggtc aggccgttga agttgaagtt 2940 cacgcatttg ttcttcacga gattggtgct tttcttaggg ccgcacactg tggcaggggc 3000 atgcagcagt tcgaagctca gcaccaccac tctgtagggc tgatagccca cgccatttgt 3060 gggctgaaag ccgtaggact gcagtgggaa gtagcagttg aagccttcca cgccgttaca 3120 aggggtgctg ccggcctgat agatctcggt ggagatgtcc cgctcgaagg gcttcagatt 3180 ggacttccgg aacagccggt acaggtaatt gtagttgccg ccgactttgg agtccaggtt 3240 gttgctgttc caggcaatca cacagccggt gaagtcgtcg ggcagcttgt agttgtagtc 3300 ggcgatcttg cctgtctgtc caggggcaat ctgccgcact tcatctcccc ggatcacgaa 3360 gctgtcggcg tacacgtttg tgaagcacag gtcgttcagc ttggtagggg acacgccgta 3420 gcacttgaag gtgctgaagc tggcggagtt gtacagcacg gagtagtcgg ccacgcaatt 3480 gctgatccgc ttccggttcc aggcgtacac agaggcgaat ctggtggcat tgaacacctc 3540 gccgaagggg cacagattgg tgatattggg gaaccgcacg atggattcgg tgggctgcac 3600 ccggaagttg ctggtctggt agatgccctt ttccacggtg aaggacttca gggtgcactt 3660 tgtctcgctc agaggatcca gagcacaatc cacggcgtcg gtgatggtgc cgttctcgtt 3720 gtacttcagc aggaaggttc taggctgcag gtagcccaca tagtaagcgg cggcaccagc 3780 tgtccatccg ctgctgctat cgccaggtgt caggtagctt ctgtgcaggg ccagcagtgt 3840 ctgaaaccgg gtgatgttga tgccgatggg cagatccacc aggggttcca gagcagagaa 3900 gccctgaggc agatcccgca cgaggttgat aggggtgtgc ttgctgtaga tcttgaagta 3960 gccgtcgatg ttcttgaaca cgaactcgcg caggttcttg aagttgccct gcttgccttc 4020 caggtccatc aggaaaggct gggacacgta ctcgaaggtg cagttgttgg cgctgctgta 4080 cacccggaac tcgctttcca tccagctctt gttgttcttg tgatagtaga cgcccaggaa 4140 ggggtcgttg cagaactgga actcgcacac tttgatgacc acgttggtgg cgttgttcac 4200 gatcagcagg ctctgggtct tgctgtccag tgtggtgccg aagatccagc ctctgatgat 4260 gttggacttc tcggtgctgg caaagtacac cccgtcgttg aagggcagca cggggttgtc 4320 gaatctcttg gtgccattgg tgccggacac gtggatggcg tggaaccagg tcacgttgct 4380 gaaaaggc aggaacaggt cctgggtaga gtgcagcacg ctggatctga acaccttgtc 4440 ggggtagtac acgcctctgg taaagctgtt ggtgtaggct ggaggcagct gggttcttgt 4500 ggtcaggttc acacactgac tagagactag tggcaataac acaagcacaaacatggt 4560 accaagctta ggctcgagcg gccgcgtcga cggtaccaga tctctagcgg atctgacggt 4620 tcactaacc agctctgctt atatagacct cccaccgtac acgcctaccg cccatttgcg 4680 tcaatggggc ggagttgtta cgacattttg gaaagtcccg ttgattttgg tgccaaaaca 4740 aactcccatt gacgtcaatg gggtggagac ttggaaatcc ccgtgagtca aaccgctatc 4800 cacgcccatt gatgtactgc caaaaccgca tcaccatggt aatagcgatg actaatacgt 4860 agatgtactg ccaagtagga aagtcccata aggtcatgta ctgggcataa tgccaggcgg 4920 gccatttacc gtcattgacg tcaatagggg gcgtacttgg catatgatac acttgatgta 4980 ctgccaagtg ggcagtttac cgtaaatact ccacccattg acgtcaatgg aaagtcccta 5040 ttggcgttac tatgggaaca tacgtcatta ttgacgtcaa tgggcgggg tcgttgggcg 5100 gtcagccagg cgggccattt accgtaagtt atgtaacgcg gaactccata tatgggctat 5160 gaactaatga ccccgtaatt gattactatt aacagtgttt aaacgtatac ctataaaggc 5220 gggtgtctta cgagggtctt tttgcttttc tgcagacatc atgaacggga ctggcggggc 5280 cttcgaaggg gggcttttta gcccttattt gacaacccgc ctgccgggat gggccggagt 5340 tcgtcagaat gtgatgggat cgacggtgga tgggcgccca gtgcttccag caaattcctc 5400 gaccatgacc tacgcgaccg tggggaactc gtcgctcgac agcaccgccg cagccgcggc 5460 agccgcagcc gccatgacag cgacgagact ggcctcgagc tacatgccca gcagcggtag 5520 tagcccctct gtgcccagtt ccatcatcgc cgaggagaaa ctgctggccc tgctggccga 5580 gctgggaagcc ctgagccgcc agctggccgc cctgacccag caggtgtccg agctccgcga 5640 acagcagcag cagcaaaata aatgattcaa taaacacaga ttctgattca aacagcaaag 5700 catctttat atttatttt tcgcgcgcgg taggccctgg tccacctctc ccgatcattg 5760 agagtgcggt ggattttttc caggacccgg tagaggtggg attggatgtt gaggtacatg 5820 ggcatgagcc cgtcccgtgg gtggaggtag caccactgca tggcctcgtg ctctggggtc 5880 gtgttgtag tgatccagtc atagcagggg cgctgggcgt ggtgctggat gatgtccttg 5940 aggaggac tgatggccac ggggagcccc ttggtgtagg tgttggcaaa acggttgagc tgggagggat gcatgcgggg ggagatgatg tgcagtttgg cctggatctt gaggttggcg 6060 atgttgccac ccagatcccg ccggggggttc atgttgtgca ggaccaccag aacggtgtag cccgtgcact tggggaactt gtcatgcaac ttggaagga atgcgtggaa gaatttggag accccttgt gcccgcccag gttttccatg cactcatcca tgatgatggc aatgggcccg 6240. tgggctgcgg ctttggcaaa gacgtttctg gggtcagaga catcgtaatt atgctcctgg gtgagatcat cataagacat tttaatgaat ttggggcgga gggtgccaga ttgggggacg atggttccct cgggccccgg ggcgaagttc ccctcgcaga tctgcatctc ccaggctttc 6420. atctcggagg gggggatcat gtccacctgc ggggcgatga aaaaaacggt ttccggggcg 6480. ggggtgatga gctgcgagga gagcaggttt ctcaacagct gggacttgcc gcacccggtc 6540 gggccgtaga tgaccccgat gacgggttgc aggtggtagt tcaaggacat gcagctgccg 6600 tcgtcccgga ggaggggggc cacctcgttg agcttgtctc tgacttggag gttttcccgg 6660 acgagctcgc cgaggaggcg gtccccgccc agcgagagaa gctcttgcag ggaagcaaag 6720 tttttcaggg gcttgagccc gtcggccatg ggcatcttgg cgagggtctg cgagaggagc 6780 tccaggcggt cccagagctc ggtgacgtgc tctacggcat ctcgatccag cagacttcct 6840 cgtttcgggg gttgggacga ctgcgactgt agggcacgag acgatgggcg tccagcgcgg 6900 ccagcgtcat gtccttccag ggtctcaggg tccgcgtgag ggtggtctcc gtcacggtga 6960 aggggtgggc cgcgggctgg gcgcttgcaa gggtgcgctt gagactcatc ctgctggtgc 7020 tgaaacgggc acggtcttcg ccctgcgcgt cggcgagata gcagttgacc atgagctcgt 7080 agttgagggc ctcggcggcg tggcccttgg cgcggagctt gcccttggaa gagcgcccgc 7140 aggcgggaca gaggagggat tgcagggcgt agagcttggg cgcgagaaag acggactcgg 7200 gggcgaaggc gtccgctccg cagtgggcgc agacggtctc gcactcgact agccaggtga 7260 gctcgggctg ctcggggtca aaaaccagtt ttcccccgtt cttttgatg cgcttcttac 7320 ctcgcgtctc catgagtctg tgtccgcgct cggtgaaaa caggctgtct gtgtccccgt 7380 agacggactt gatgggcctg tcctgcaggg gcgtcccgcg gtcctcctcg tagagaaact 7440 cagaccactc tgagacgaag gcgcgcgtcc acgccaagac aaaggaggcc acgtgcgagg 7500 ggtagcggtc gttgtccacc aggggtcca ccttttccac ggtatgcagg cacatgtccc 7560 cctcctccgc atccaagaag gtgattggct tgtaggtgta ggccacgtga cctggggttc 7620 ccgacgggg ggtataaaag ggggcgggtc tgtgctcgtc ctcactctct tccgcgtcgc 7680 tgtccacgag cgccagctgt tggggtaggt attccctctc aagagcgggc atgacctcgg 7740 cactcaggtt gtcagtttct agaaacgagg aggatttgat gtgggcctgc cctgccgcga 7800 tgctttttag gagactttca tccatctggt cagaaaagac tatttttta ttgtcaagct 7860 tggtggcgaa ggagccatag agggcgtttg agagaagctt ggcgatggat ctcatggtct 7920 gatttttgtc acggtcggcg cgctccttgg ccgcgatgtt gagctggaca tattcgcgcg 7980 cgacacactt ccattcgggg aagacggtgg tgcgctcgtc gggcacgatc ctgacgcgcc 8040 agccgcggtt atgcagggtg accaggtcca cgctggtggc cacctcgccg cgcaggggct 8100 cgttggtcca gcagagtctg ccgcccttgc gcgagcagaa cgggggcagc acatcaagca 8160 gatgctcgtc aggggggtcc gcatcgatgg tgaagatgcc cggacagagt tccttgtcaa 8220 aataatcgat ttttgaggat gcatcgtcca aggccatctg ccactcgcgg gcggccagcg 8280 ctcgctcgta ggggttgagg ggcggacccc aaggcatggg atgcgtgagg gcggaggcgt 8340 acatgccgca gatgtcatag acatagatgg gctccgagag gatgccgatg taggtgggat 8400 agcagcgccc cccgcggatg cttgcgcgca cgtagtcata caactcgtgc gagggggcca 8460 agaaggcggg gccgagattg gtgcgctggg gctgctcggc gcggaagacg atctggcgaa 8520 agatggcgtg cgagttggag gagatggtgg gccgttggaa gatgttaaag tgggcgtgag 8580 gcaggcggac cgagtcgcgg atgaagtgcg cgtaggagtc ttgcagcttg gcgacgagct 8640 cggcggtgac gaggacgtcc atggcgcagt agtccagcgt ttcgcggatg atgtcataac 8700 tcgcctctcc tttcttctcc cacagctcgc ggttgagggc gtattcctcg tcatccttcc 8760 agtactcccg gagcgggaat cctcgatcgt ccgcacggta agagcccagc atgtagaaat 8820 ggttcacggc cttgtaggga cagcagccct tctccacggg gagggcgtaa gcttgagcgg 8880 ccttgcggag cgaggtgtgc gtcagggcaa aggtgtccct gaccatgact ttcaagaact 8940 ggtacttgaa gtccgagtcg tcgcagccgc cgtgctccca gagctcgaaa tcggtgcgct 9000 tcttcgagag ggggttaggc agagcgaaag tgacgtcatt gaagagaatc ttgcctgccc 9060 gcggcatgaa attgcgggtg atgcggaaag ggcccgggac ggaggctcgg ttgttgatga 9120 cctgggcggc gaggacgatc tcgtcaaagc cgttgatgtt gtgcccgacg atgtagagtt 9180 ccatgaatcg cgggcggcct ttgatgtgcg gcagctttt gagctcctcg taggtgaggt 9240 cctcggggca ttgcaggccg tgctgctcga gcgccccactc ctggagatgt gggttggctt 9300 gcatgaagga agcccagagc tcgcgggcca tgagggtctg gagctcgtcg cgaaagaggc 9360 ggaactgctg gcccacggcc atcttttctg gggtgacgca gtagaaggtg agggggtccc 9420 gctcccagcg atcccagcgt aaacgcacgg cgagatcgcg agcgagggcg accagctctg 9480 ggtccccgga gaatttcatg accagcatga aggggacgag ctgcttgccg aaggacccca 9540 tccaggtgta ggtttctaca tcgtaggtga caaagagccg ctccgtgcga ggatgagagc cgattggga gaactggatt tcctgccacc agttggacga gtggctgttg atgtgatgaa agtagaaatc ccgccggcga accgagcact cgtgctgatg cttgtaaag cgtccgcagt actcgcagcg ctgcacgggc tgtacctcat ccacgagata cacagcgcgt cccttgagga 9780 ggaacttcag gagtggcggc cctggctggt ggttttcatg ttcgcctgcg tgggactcac 9840 cctggggctc ctcgaggacg gagaggctga cgagcccgcg cgggagccag gtccagatct 9900 cggcgcggcg ggggcggaga gcgaagacga gggcgcgcag ttgggagctg tccatggtgt 9960 cgcggagatc caggtccggg ggcagggttc tgaggttgac ctcgtagagg cgggtgaggg 10020 cgtgcttgag atgcagatgg tacttgattt ctacgggtga gttggtggtc gtgtccacgc 10080. attgcatgag cccgtagctg cgcggggcca cgaccgtgcc gcggtgcgct tttagaagcg gtgtcgcgga cgcgctcccg gcggcagcgg cggttccggc cccgcgggca ggggcggcag 10200 aggcacgtcg gcgtggcgct cgggcaggtc ccggtgctgc gccctgagag cgctggcgtg 10260 cgcgacgacg cggcggttga catcctggat ctgccgcctc tgcgtgaaga ccacgggccc 10320 cgtgactttg aacctgaaag acagttcaac agaatcaatc tctgcgtcat tgacggcggc 10380 ctgacgcagg atctcttgca cgtcgcccga gttgtcctgg taggcgatct cggacatgaa 10440 ctgttcgatc tcctctcct ggagatcgcc gcggcccgcg cgctccacgg tggcggcgag 10500 gtcattggag atgcgaccca tgagctgcga gaaggcgccc aggccgctct cgttccagac 10560 gcgggcttag accacgtccc cgtcggcgtc gcgcgcgcgc atgaccacct gcgcgaggtt 10620 gagctccacg tgccgcgcaa agacggcgta gttgcgcagg cgctggaaga ggtagttgag 10680 ggtggtggcg atgtgctcgg tgacgaagaa gtacatgatc cagcggcgca ggggcatctc 10740 gctgatgtcg ccgatggctt ccagcctttc catggcctcg tagaagtcca cggcgaagtt 10800 gaaaaactgg gcgttgcggg ccgagaccgt gagctcgtct tccaggagcc ggatgagttc 10860 ggcgatggtg gcgcgcacct cgcgctcgaa atccccgggg gcctcctcct cttcctcttc 10920 ttccatgacg acctcttctt ctattcttc ctctggggggc ggtggtggtg gcggggggcg 10980. acgacgacgg cgacgcaccg ggagacggtc gacgagcgc tcgatcatct ccccgcggcg 11040 gcgacgcatg gtttcggtga cggcgcgacc ccgttcgcga ggacgcagcg tgaagacgcc gccggtcatc tcccggtaat ggggcgggtc cccattgggc agcgataggg cgctgacgat 11160 gcatcttatc aattgcggtg taggggacgt gagcgcgtcg agatcgaccg gatcggagaa 11220 tctttcgagg aaagcgtcta gccaatcgca gtcgcaaggt aagctcaaac acgtagcagc cctgcggacg ctgttagaat tgcggttgct gatgatgtaa ttgaagtagg cgtttttgag gcggcggatg gtggcgagga ggaccaggtc cttgggtcca gcttgctgga tgcggagccg 11400 ctcggccatg ccccaggcct ggccctgaca ccggctcagg ttcttgtagt agtcatgcat 11460 gagcctctca atgtcatcac tggctgaggc ggagtcttcc atgcgggtga ccccgacgcc 11520. cctgagcggc tgcacgagcg ccaggtcggc gacgacgcgc tcggcgagga tggcctgttg 11580 cacgcgggtg agggtgtcct ggaagtcgtc catgtcgacg aagcggtgat aggccccggt 11640 gttgatggtg taggtgcagt tggccatgag cgaccagttg acggtctgca ggcctggctg 11700 cacgacctcg gagtacctga gccgcgagaa ggcgcgcgag tcgaagacgt agtcgttgca 11760 ggtgcgcacg aggtactggt atccgactag gaagtgcggc ggcggctggc ggtagagcgg 11820 ccagcgctgg gtggccggcg cgcccggggc caggtcctcg agcatgaggc ggtggtagcc 11880 gtagaggtag cgggacatcc aggtgatgcc ggcggcggtg gtggaggcgc gcgggaactc 11940 gcggacgcgg ttccagatgt tgcgcagcgg caggaaatag tccatggtcg gcacggtctg 12000 gccggtgaga cgcgcgcagt cattgacgct ctagaggcaa aaacgaaagc ggttgagcgg 12060 gctcttcctc cgtagcctgg cggaacgcaa acgggttagg ccgcgtgtgt accccggttc 12120 gagtcccctc gaatcaggct ggagccgcga ctaacgtggt attggcactc ccgtctcgac 12180 ccgagcccga tagccgccag gatacggcgg agagcccttt ttgctggccg aggggggtcg 12240 ctagacttga aagcgaccga aaaccctgcc gggtagtggc tcgcgcccgt agtctggaga 12300 agcatcgcca gggttgagtc gcggcagaac ccggttcgag gacggccgcg gcgagcggga 12360 cttggtcacc ccgccgatat aaagacccac agccagccga cttctccagt tacgggagcg 12420 agcccccttt tttctttttg ccagatgcat cccgtcctgc gccaaatgcg tcccaccccc 12480 ccggcgacca ccgcgaccgc ggccgtagca ggcgccggcg ctagccagcc accacagaca 12540 gagatggact tggaagaggg cgaagggctg gcaagactgg gggcgccgtc cccggagcga 12600 catccccgcg tgcagctgca gaaggacgtg cgcccggcgt acgtgcctac gcagaacctg 12660 ttcagggacc gcagcgggga ggagcccgag gagatgcgcg actgccggtt tcgggcgggc 12720 agggagctgc gcgagggcct ggaccgccag cgcgtgctgc gcgacgagga tttcgagccg 12780 aacgagcaga cggggatcag ccccgcacgc gcgcacgtgg cggcagccaa cctggtgacg 12840 gcctacgagc agacggtgaa gcaggagcgc aacttccaaa agagtttcaa caaccacgtg 12900 cgcaccctga tcgcgcgcga ggaggtggcc ctgggcctga tgcacctgtg ggacctggcg 12960 gaggccatcg tgcagaaccc ggacagcaag cctctgacgg cgcagctgtt cctggtggtg cagcacagca gggacaacga ggcgttcagg gaggcgctgc tgaacatcgc cgagcccgag ggtcgctggc tgctggagct gattaacatc ttgcagagca tcgtagtgca ggagcgcagc ctgagcctgg ccgagaaggt ggcggcgatc aactactcgg tgctgagcct gggcaagttt 13200 tacgcgcgca agatttacaa gacgccgtac gtgcccatag acaaggaggt gaagatagac agcttttaca tgcgcatggc gctcaaggtg ctgacgctga gcgacgacct gggcgtgtac cgcaacgacc gcatccacaa ggccgtgagc acgagccggc ggcgcgagct aagcgaccgc gagctgatgc tgagtctgcg ccggggcgctg gtagggggcg ccgccggcgg cgaggagtcc 13440 tacttcgaca tgggtgcgga cctgcattgg cagccgagcc ggcgcgcctt ggaggccgcc 13500 tacggttcag aggacttgga tgaggaagag gaagaggagg aggatgcacc cgctgcgggg 13560 tactgacgcc tccgtgatgt gtttttagat gtcccagcaa gccccggacc ccgccataag ggcggcgctg caaagccagc cgtccggtct agcatcggac gactgggagg ccgcgatgca 13680 acgcatcatg gccctgacga cccgcaaccc cgagtccttt agacaacagc cgcaggccaa 13740 cagactctcg gccattctgg aggcggtggt cccctctcgg accaacccca cgcacgagaa 13800 ggtgctggcg atcgtgaacg cgctggcgga gaacaaggcc atccgtcccg acgaggccgg 13860 gctggtgtac aacgccctgc tggagcgcgt gggccgctac aacagcacga acgtgcagtc 13920 caacctggat cggctggtga cggacgtgcg cgaggccgtg gcgcagcgcg agcggttcaa 13980 gaacgagggc ctgggctcgc tggtggcgct gaacgccttc ctggcaacgc agccggcgaa 14040 cgtgccgcgc gggcaggacg attacaccaa ctttatcagc gcgctgcggc tgatggtgac 14100 cgaggtgccc cagagcgagg tgtaccagtc tggcccggac tactttttcc agacgagccg 14160 gcagggcttg cagacggtga acctgagcca ggctttcaag aatctgcgcg ggctgtgggg 14220 cgtgcaggcg cccgtgggcg accggtcaac ggtgagcagc ttgctgacgc ccaactcgcg 14280 gctgctgctg ctgctgatcg cgcccttcac cgacagcggc agcgtgaacc gcaactcgta 14340 cctgggccat ctgctgacgc tgtaccgcga ggccataggc caggcgcagg tggacgagca 14400 gaccttccag gagatcacta gcgtgagccg cgcgctgggg cagaacgaca ccgacagtct 14460 gagggccacc ctgaactttt tgctgaccaa tagacagcag aagatcccgg cgcagtacgc 14520 actgtcggcc gaggaggaaa ggattctgag atatgtgcag cagagcgtag ggctgttcct 14580 gatgcaggag ggtgccaccc ccagcgccgc gctggacatg accgcgcgca acatggaacc 14640 tagcatgtac gccgccaacc ggccgttcat caataagctg atggactact tgcaccgcgc 14700 ggcggccatg aacacggact actttaccaa cgccatcctg aacccgcact ggctcccgcc 14760 gccggggttc tacacgggcg agtacgacat gcccgacccc aacgacgggt tcctgtggga 14820 cgacgtggac agcgcggtgt tctcgccgac ctttcaaaag cgccaggagg cgccgccgag 14880 cgagggcgcg gtggggagga gcccctttcc tagcttaggg agtttgcata gcttgccggg 14940 ctcggtgaac agcggcaggg tgagccggcc gcgcttgctg ggcgaggacg agtacctgaa 15000 cgactcgctg ctgcagccgc cgcgggccaa gaacgccatg gccaataacg ggatagagag 15060 tctggtggac aaactgaacc gctggaagac ctacgctcag gaccataggg acgcgcccgc 15120 gccgcggcga cagcgccacg accggcagcg gggcctggtg tgggacgacg aggactcggc 15180 cgacgatagc agcgtgttgg acttgggcgg gagcggtggg gtcaacccgt tcgcgcatct 15240 gcagcccaaa ctggggcgac ggatgttttg aatgaaataa aactcaccaa ggccatagcg 15300 tgcgttctct tccttgttag agatgaggcg cgcggtggtg tcttcctctc ctcctccctc 15360 gtacgagagc gtgatggcgc aggcgaccct ggaggttccg tttgtgcctc cgcggtatat 15420 ggctcctacg gagggcagaa acagcattcg ttactcggag ctggctccgc agtacgacac 15480 cactcgcgtg tacttggtgg acaacaagtc ggcggacatc gcttccctga actaccaaaa 15540 cgaccacagc aacttcctga ccacggtggt gcagaacaac gatttcaccc ccgccgaggc 15600 cagcacgcag acgataaatt ttgacgagcg gtcgcggtgg ggcggtgatc tgaagaccat 15660 tctgcacact aacatgccca atgtgaacga gtacatgttc accagcaagt ttaaggcgcg 15720 ggtgatggtg tctaggaagc atccagaggg ggtagttgaa acagatttga gtcaggataa 15780 gcttgaatat gagtggtttg agtttaccct gcccgaggga aacttttccg agaccatgac 15840 15900 cgtgctggag agcgatatcg gagtcaagtt tgacagcaga aatttcaagc tgggctggga 15960 cccggtgacc aagctggtga tgccaggggt ctacacctac gaggccttcc acccggacgt 16020 ggtgctgctg ccgggctgcg gggtggactt caccgagagc cgcctgagca acctcctggg 16080 cattcgcaag aagcaacctt tccaagaggg cttcagaatc atgtatgagg atctagaagg 16140 16200 agaacgaact aaaaatgcag ctgcggccac agccgataca accactaggg gtgatacatt 16260 tgcaactcca gcgcaagaga cagcagctga taagaggta gaagtcttgc ccattgaaaa 16320 ggatgagagt ggtagaagtt acaacctgat ccaggggacc cacgacacgc tgtaccgcag 16380 ttggtacctg tcctatacct acggggaccc cgagaagggg gtgcagtcgt ggacgctgct 16440 caccaccccg gacgttacct gcggcgcgga gcaagtctac tggtcactgc cggacctcat 16500 gcaagacccc gtcaccttcc gctccaccca gcaagtcagc aactaccccg tggtcggcgc 16560 cgagctcatg cccttccgcg ccaagagctt ttacaacgac ctcgccgtct actcccagct 16620 catccgcagc tacacctccc tcacccacgt cttcaaccgc ttccccgaca accagatcct 16680 ctgccgcccg cccgcgccca ccatcaccac cgtcagtgaa aacgtgcctg ctctcacaga 16740 tcacgggacg ctaccgctgc gcagcagtat ccgcggagtc cagcgagtga ccgtcactga 16800 cgcccgtcgc cgcacctgtc cctacgtcta caaggccctg ggcatagtcg cgccgcgcgt 16860 gctttccagt cgcaccttct aaaaaaatgt ctattctcat ctcgcccagc aataacaccg 16920 gctggggtct tactagaccc agcaccatgt acggaggagc caagaagcgc tcccagcagc 16980 accccgtccg cgtccgcggc cacttccgcg ctccctgggg cgcttacaag cgcgggcgga 17040 cttcaccgc cgtgcgcacc accgtcgacg acgtcatcga ctcggtggtc gccgacgcgc 17100 gcaactacac tcccgcccc tccaccgtgg acgcggtcat cgacagcgtg gtggccgacg 17160 cgcgcgacta tgccagacgc aagagccggc ggcgacggat cgccaggcgc caccggagca 17220 cgcccgccat gcgcgccgcc cgggctctgc tgcgccgcgc cagacgcacg ggccgccggg 17280 ccatgatgcg agccgcgcgc cgcgctgcca ctgcacccac ccccgcaggc aggactcgca 17340 gacgagcggc cgccgccgcc gctgcggcca tctctagcat gaccagaccc aggcgcggaa 17400 acgtgtactg ggtgcgcgac tccgtcacgg gcgtgcgcgt gcccgtgcgc acccgtcctc 17460 ctcgtccctg atctaatgct tgtgtcctcc cccgcaagcg acgatgtcaa agcgcaaaat 17520 caaggaggag atgctccagg tcgtcgcccc ggagagatttac ggaccacccc aggcggacca 17580 gaaccccgc aaaatcaagc gggtttaaaaa aaaggatgag gtggacgagg gggcagtaga 17640 gtttgtgcgc gagttcgctc cgcggcggcg cgtaaattgg aaggggcgca gggtgcagcg 17700 cgtgttgcgg cccggcacgg cggtggtgtt cacgccccggc gagcggtcct cggtcaggag 17760 caagcgtagc tatgacgagg tgtacggcga cgacgacatc ctggaccagg cggcggagcg 17820 ggcgggcgag ttcgcctacg ggaagcggtc gcgcgaagg gagctgat cgctgccgct 17880 ggacgaaagc aaccccacgc cgagcctgaa gcccgtgacc ctgcagcagg tgctgcccca 17940 ggcggtgctg ctgccgagcc gcggggtcaa gcgcgaggggc gagagcatgt acccgaccat 18000 gcagatcatg gtgcccaagc gccggcgcgt ggaggacgtg ctggacaccg tgaaaatgga 18060 tgtggagccc gaggtcagg tgcgcccat caagcaggtg gcgccgggcc tgggcgtgca 18120 aaccgtggac attcagatcc cccacccat ggatgtcgac aaaaaaccct cgaccagcat 18180 cgaggtgcaa accgaccct ggctcccagc ctccaccgct accgtctcca cttctaccgc 18240 cgccacggct accgagccctc ccaggaggcg aagatggggc gccgccagcc ggctgatgcc 18300 caacacgtg ttgcatcctt ccatcatccc gacgccgggc taccgcggca cccgtacta 18360 cgccagccgc cggcgcccag ccagcaacg ccgccgccgc accgccc gccgccgtct 18420 ggcccccgcc cgcgtgcgcc gcgtgaccac gcgccggggc cgctcgctcg ttctgcccac 18480 cgtgcgctac caccccagca tcctttaatt cgtgtgctgt gatactgttg cagagagatg 18540 gctctcactt gccgcctgcg catccccgtc ccgaattacc gaggaagatc ccgccgcagg 18600 agaggcatgg caggcagcgg cctgaaccgc cgccggcggc gggccatgcg caggcgcctg 18660 agtggcggct ttctgcccgc gctcatcccc ataatcgccg cggccattgg cacgatcccg 18720 ggcatagctt ccgttgcgct gcaggcgtcg cagcgccgtt gatgtgcgaa taaagcctct 18780 ttagactctg acacacctgg tcctgtatat ttttagaatg gaagacatca attttgcgtc 18840 cctggctccg cggcacggca cgcggccgtt catgggcacc tggaacgaga tcggcaccag 18900 ccagctgaac gggggcgcct tcaattggag cagtgtctgg agcgggctta aaaatttcgg 18960 ctcgacgctc cggacctatg ggaacaaggc ctggaatagt agcacggggc agttgttaag 19020 ggaaaagctc aaagaccaaa acttccagca gaaggtggtg gacgggctgg cctcgggcat 19080 taacggggtg gtggacatcg cgaaccaggc cgtgcagcgc gagataaaca gccgcctgga 19140 cccgcggccg cccacggtgg tggagatgga agatgcaact cttccgccgc ccaaaggcga 19200 aaagcggccg cggcccgacg cggaggagac gatcctgcag gtggacgagc cgccctcgta 19260 cgaggaggcc gtcaaggccg gcatgcccac cacgcgcatc atcgcgccgc tggccacggg 19320 tgtaatgaaa cccgccaccc ttgacctgcc tccaccaccc gcgcccgctc caccgaaggc 19380 aactccggtt gtgcaggccc ccccggtggc gaccgccgtg cgccgcgtcc ccgcccgccg 19440 ccaggcccag aactggcaga gcacgctgca cagtatcgtg ggcctgggag tgaaaagtct 19500 gaagcgccgc cgatgctatt gagagagagg aaagaggaca ctaaagggag agcttaactt 19560 gtatgtgcct taccgccaga gaacgcgcga agatggccac cccctcgatg atgccgcagt 19620 gggcgtacat gcacatcgcc gggcaggacg cctcggagta cctgagcccg ggtctggtgc 19680 agtttgcccg cgccaccgac acgtacttca gcctgggcaa caagtttagg aaccccacgg 19740 tggccccgac ccacgatgtg accacggacc ggtcccagcg tctgacgctg cgcttcgtgc 19800 ccgtggatcg cgaggacacc acgtactcgt acaaggcgcg cttcactctg gccgtgggcg 19860 acaaccgggt gctagacatg gccagcactt actttgacat ccgcggcgtc ctggaccgcg gtcccagctt caaaccctac tcgggcacgg cctacaacg cctggctccc aagggtgccc 19980. ccaatcccag tcagtggga acaaaagaaa agcaaggaac tactggagga gtgcagcaag aaaagatgt cacaaaaaca tttggtgtgg ctgccaccgg cggaattaat ataacaaacc agggtctgtt actaggaact gacgaaccg ctgagaatgg caaaaaagac atttatgcag acaagacttt ccagccaga cctcaagttg gagaagaaaa ctggcagga aatgaagcct tctatggagg aagggctctt aaaaaggaca ctaaaatgaa accatgctat ggatcttttg ctagacctac taatgagaaa ggaggtcagg caaagttcaa accagttaat gaggagac aacctaaaga tctggatata gattttgctt actttgacgt ccctggcgga agtcctccag caggtggtag tggggaga tacaaagcag atataatttt gtacactgaa aatgttaatc ttgaaacacc agacactcat gtggtttaca agccaggaac ttcagataac agttcagaaa tcaatctggt tcagcagtcc atgccaaaca gacccaacta cattggcttt agggacaact 20580 ttgtaggtct catgtattac aacagcaccg gaaatatggg tgtgctggct ggtcaggctt 20640 ctcagttgaa cgctgtggtc gacttgcaag acagaaacac cgagttatct taccagctat 20700 tgctagattc tctgggtgac agaaccagat actttagcat gtggaactct gcggtggaca 20760 gttacgatcc agatgtcagg atcattgaaa atcacggtgt ggaagatgaa cttccaaact 20820 attgcttccc attgaatggc actggaacca attccactta tcaaggtgta aagattacaa 20880 atggtaatga tggtgctgaa gaaagtgagt gggagaaaga cgatgcaatt tctagacaaa 20940 accaaatctg caagggcaat gtctacgcca tggagatcaa cctgcaggcc aacctgtgga 21000 agagttttct gtactcgaac gtggccctgt acctgcccga ctcctacaag tacacgccgg 21060 ccaacgtcaa gctgcccgcc aacaccaaca cctacgagta catgaacggc cgcgtggtag 21120 ccccatccct ggtggacgcc tacatcaaca tcggcgcccg ctggtcgttg gaccccatgg 21180 acaacgtcaa ccccttcaac caccaccgca atgcgggcct gcgctaccgc tccatgctgc 21240 tgggcaacgg ccgctacgtg cccttccaca tccaagtgcc ccaaaagttc tttgccatca 21300 agaacctgct cctgctcccg ggctcctaca cctacgagtg gaacttccgc aaggacgtca 21360 acatgatcct gcagagttcc ctcggcaacg acctgcgcgt cgacggcgcc tccgtccgct 21420 tcgacagcgt caacctatac gccactttct tccccatggc gcacaacacc gcttcaacct 21480 tggaagccat gctgcgcaac gacaccaacg accagtcctt caacgactac ctctcggccg 21540 ccaacatgct ctaccccatc ccggccaagg ccaccaacgt gcccatctcc atcccatcgc 21600 gcaactgggc cgccttccgc ggctggagtt tcacccggct caagaccaag gaaactcctt 21660 ccctcggctc gggtttcgac ccctactttg tctactcggg ctccatcccc tacctcgacg 21720 ggaccttcta cctcaaccac accttcaaga aggtctccat catgttcgac tcctcggtca 21780 gctggcccgg caacgaccgg ctgctcacgc cgaacgagtt cgagatcaag cgcagcgtcg 21840 acggggaggg ctacaacgtg gcccaatgca acatgaccaa ggactggttc ctcgtccaga 21900 tgctctccca ctacaacatc ggctaccagg gcttccacgt gcccgagggc tacaaggacc 21960 gcatgtactc cttcttccgc aacttccagc ccatgagcag gcaggtggtc gatgagatca 22020 actacaagga ctacaaggcc gtcaccctgc ccttccagca caataactcg ggcttcaccg 22080 gctacctcgc acccaccatg cgccaggggc agccctaccc cgccaacttc ccctacccgc 22140 tcatcggtca gacagccgtg ccctccgtca cccagaaaaa gttcctctgc gacagggtca 22200 tgtggcgcat cccattctcc agcaacttca tgtccatggg cgccctcacc gacctgggtc 22260 agaacatgct ctacgccaac tcggcccacg cgctcgacat gaccttcgag gtggacccca 22320 tggatgagcc caccctcctc tatcttctct tcgaagtttt cgacgtggtc agagtacacc 22380 agccgcaccg cggcgtcatc gaggccgtct acctgcgcac gcccttctcc gccggcaacg 22440 ccaccaccta agcatgagcg gctccagcga acgagagctc gcggccatcg tgcgcgacct 22500 gggctgcggg ccctactttt tgggcaccca cgacaagcgc ttcccgggct ttctcgccgg 22560 cgacaagctg gcctgcgcca tcgtcaacac ggccggccgc gagaccggag gcgtgcactg gctcgccttc ggctggaacc cgcgctcgcg cacctgctac atgttcgacc cctttgggtt 22680 ctcggaccgc cggctcaagc agatttacag cttcgagtac gaggccatgc tgcgccgcag cgccctggcc tcctcgcccg accgctgtct cagcctcgag cagtccactc agaccgtgca 22800 ggggcccgac tccgccgcct gcggactctt ctgttgcatg ttcttgcatg ccttcgtgca 22860 ctggcccgac cgacccatgg acggaaccc caccatgac ttgctgacgg gggtgcccaa cggcatgcta caatcgccac aggtgctgcc caccctcagg cgcaaccagg aggaactcta ccgcttcctc gcgcgccact ccccttactt tcgctcccac cgcgccgcca tcgaacacgc 23040 caccgctttt gacaaaatga aacaactgcg tgtatctcaa taaacagcac ttttatttta catgcactgg agttatgca agttatttaa aagtcgaagg ggttctcgcg ctcgtcgttg 23160 tgcgccgcgc tggggagggc cacgttgcgg tactggtact tgggctgcca cttgaactcg 23220 gggatcacca gtttgggcac tggggtctcg gggaaggtct cgctccacat gcgccggctc 23280. atctgcaggg cgcccagcat gtccggggcg gagatcttga aatcgcagtt ggggccggtg 23340 ctctgcgcgc gcgagttgcg gtacacgggg ttgcagcact ggaacaccat cagactgggg 23400 tacttcacac tagccagcac gctcttgtcg ctgatctgat ccttgtccag atcctcggcg 23460 ttgctcaggc cgaacggggt catcttgcac agctggcgtc ccaggaaggg cacgctctga 23520 ggcttgtggt tacactcgca gtgcacgggc atcagcatca tccccgcgcc gcgctgcata 23580 ttcgggtaga gggccttgac aaaggccgcg atctgcttga aagcttgctg ggccttggcc 23640 ccctcgctga aaaacaggcc gcagctcttc ccgctgaact ggttattccc acacccggca 23700 tcctgcacgc agcagcgcgc gtcatggctg gtcagttgca ccacgctccg tccccagcgg 23760 ttctgggtca ccttagcctt gctgggctgc tccttcaacg cgcgctgccc gttctcgctg 23820 gtcacatcca tctccaccac gtggtccttg tggatcatca tcgtcccgtg cagacacttg 23880 agctggcctt ccacctcggt gcagccgtga tcccacaggg cgcaaccggt gcactcccag 23940 ttcttgtgcg caatcccgct gtggctgaag atgtaacctt gcaacatgcg gcccatgatg 24000 gtgctaaatg ctttctgggt ggtgaaggtc agttgcatcc cgcgggcctc ctcgttcatc 24060 caggtctggc acatcttctg gaagatctcg gtctgctcgg gcatgagctt gtaagcatcg 24120 cgcaggccgc tgtcgacgcg gtagcgttcc atcagcacgt tcatggtatc catgcccttc 24180 tcccaggacg agaccagagg cagactcaga gggttgcgta cgttcaggac accgggggtc 24240 gcgggctcga cgatgcgttt tccgtccttg ccttccttca atagaaccgg cggctggctg 24300 aatcccactc ccacgatcac ggcatcttcc tggggcatct cttcgtcggg gtctaccttg 24360 gtcacatgct tggtctttct ggcttgcttc ttttttggag ggctgtccac ggggagcacg 24420 tcctcctcgg aagacccgga gcccacccgc tgatactttc ggcgcttggt gggcagagga 24480 ggtggcggcg aggggctcct ctcctgctcc ggcggatagc gcgccgaccc gtggccccgg 24540 ggcggagtgg cctctcggcc catgaaccgg cgcacgtcct gactgccgcc ggccattgtt 24600 tcctagggga agatggagga gcagccgcgt aagcaggagc aggaggagga cttaaccacc 24660 cacgagcaac ccaaaatcga gcaggacctg ggcttcgaag agccggctcg tctagaaccc 24720 ccacaggatg aacaggagca cgagcaagac gcaggccagg aggagaccga cgctgggctc 24780 gagcatggct acctgggagg agaggaggat gtgctgctga aacacctgca gcgccagtcc 24840 ctcatcctcc gggacgccct ggccgaccgg agcgaaaccc ccctcagcgt cgaggagctg 24900 tgtcgggcct acgagctcaa cctcttctcg ccgcgcgtac cccccaaacg ccagcccaac 24960 ggcacctgcg agcccaaccc gcgtctcaac ttctatcccg tctttgcggt ccccgaagcc 25020 ctcgccacct atcacatctt tttcaagaac caaaagatcc ccgtctcctg ccgcgccaac 25080 cgcaccagcg ccgacgcgct cctcgctctg gggcccggcg cgcgcatacc tgatatcgct 25140 tccctggaag aggtgcccaa gatcttcgaa gggctcggtc gggacgagac gcgcgcggcg 25200 aacgctctga aagaaacagc agaggaagag ggtcacacta gcgccctggt agagttggaa 25260 ggcgacaacg ccaggctggc cgtgctcaag cgcagcgtcg agctcaccca cttcgcctac 25320 cccgccgtca acctcccgcc caaggtcatg cgtcgcatca tggatcagct catcatgccc 25380 cacatcgagg ccctcgatga aagtcaggag cagcgccccg aggacgcccg gcccgtggtc 25440 agcgacgaga tgctcgcgcg ctggctcggg acccacgacc cccaggcttt ggaacagcgg 25500 cgcaagctca tgctggccgt ggtcctggtt accctcgagc tggaatgcat gcgccgcttc 25560 ttcagcgacc ccgagaccct gcgcaaggtc gaggagaccc tgcactacac tttcagacac 25620 ggtttcgtca ggcaggcctg caagatctcc aacgtggagc tgaccaacct ggtctcctgc 25680 ctggggatcc tgcacgagaa ccgcctgggg cagaccgtgc tccactctac cctgaagggc 25740 gaggcgcggc gggactatgt ccgcgactgc gtctttctat ttctttgcca cacatggcaa 25800 gcagccatgg gcgtgtggca acagtgtctc gaggacgata acctgaagga gctggacaag 25860 cttcttgcta gaaatcttaa aaagctgtgg acgggcttcg acgagcgcac cgtcgcctcg 25920 gacctggccg agatcgtgtt ccccgagcgc ctgaggcaga cgctgaaagg cgggctgccc 25980 gacttcatga gccagagcat gttgcaaaac taccgcactt tcattctcga gcgatctggg 26040 atgctgcccg ccacctgcaa cgctttcccc tccgactttg tcccgctgag ctaccgcgag 26100 tgtcccccgc cgctgtggag ccactgctac ctcttgcagc tggccaacta catcgcctac 26160 cactcggacg tgatcgagga cgtgagcggc gaggggctgc tcgagtgcca ctgccgctgc 26220 aacctgtgct ccccgcaccg ctccctggtc tgcaaccccc agctactaag cgagacccag 26280 gtcatcggta cctttgagct gcaaggtccg caggagtcca ccgctccgct gaaactcacg 26340 ccggggttgt ggacttccgc gtacctgcgc aaatttgtac ccgaggacta ccacgcccac 26400 gagataaagt tcttcgagga ccaatcgcgt ccgcagcacg cggatctcac ggcctgcgtc 26460 atcacccagg gcgcaatcct cgcccaattg cacgccatcc aaaaatcccg ccaagagttt 26520 cttctgaaaa agggtagagg ggtctacctg gacccccaga cgggcgaagt gctcaacccg 26580 ggtctccccc agcatgccga ggaagaagca ggagccgcta gtggaggaga tggaagaaga 26640 atgggacagc caggcagagg aggacgaatg ggaggaggag acagaggagg aagaattgga 26700 agaggtggaa gaggagcagg cacagagca gcccgtcgcc gcaccatccg cgccggcagc 26760 cccgccggtc acggatacaa cctccgcagc tccggccaag cctcctcgta gatgggatcg 26820 agtgaagggt gacggtaagc acgagcggca gggctaccga tcatggagggg cccacaagc 26880 cgcgatcatc gcctgcttgc aagactgcgg gggaacatc gcttcgccc gccgctacct 26940 gctcttccac cgcggggtaa acatcccccg caacgtgttg cattactacc gtcaccttca 27000 cagctaagaa aaagcaagta aaaggagtcg ccggaggagg aggaggcctg aggatcgcgg 27060 cgaacgagcc cttgaccacc agggagctga ggaaccggat cttccccact ctttatgcca 27120 ttttcagca gagtcgaggt cagcagcaag agctcaagt aaaaaccgg tctctgcgct 27180 cgctcacccg cagttgcttg taccacaaa acgagatca gctgcagcgc actctcgaag 27240 acgccgaggc tctgttccac aagtactgcg cgctcactct taagacta ggcgcgccca 27300 cccggaaaaa agggggaat tacctcatcg cccacatgag cagggatt cccacccctt 27360 acatgtggag ctatcagccc caatgggcc tggccgcggg cgcctcccag gactactcca 27420 cccgcatgaa ctggctcagt gccggcccct cgatgatctc acgggtcaac ggggtccgca 27540. gtcatcgaaa ccagatattg ttggagcagg cggcggtcac ctccacgccc agggcaaagc tcaacccgcg taattggccc tccaccctgg tgtatcagga aatccccggg ccgactaccg tactacttcc gcgtgacgca ctggccgaag tccgcatgac taactcaggt gtccagctgg ccggcggcgc ttcccggtgc ccgctccgcc cacaatcggg tataaaaacc ctggtgatcc gaggcagagg cacacagctc aacgacgagt tggtgagctc ttcgatcggt ctgcgaccgg acggagtgtt ccaactagcc ggagccgggga gatcctcctt cactcccaac caggcctacc tgaccttgca gagcagctct tcggagcctc gctccggagg catcggacc ctccagtttg tggaggagtt tgtgccctcg gtctacttca accccttctc gggatcgcca ggcctctacc 27960 cggacgagtt cataccgac ttcgacgcag tgagagagc ggtggacggc tacgactgaa tgtcccatgg tgactcggct gagctcgctc ggttgaggca tctggaccac tgccgccgcc 28080 tgcgctgctt cgcccgggag agctgcggac tcatctactt tgagtttccc gaggagcacc 28140 ccaacggccc tgcacacgga gtgcggatca ccgtagaggg caccaccgag tctcacctgg 28200 tcaggttctt cacccagcaa cccttcctgg tcgagcggga ccggggcgcc accacctaca 28260 ccgtctactg catctgtcca accccgaagt tgcatgagaa tttttgttgt actctttgtg 28320 gtgagtttaa taaaagctaa actcttgcaa tactctggac cttgtcgtcg tcaactcaac 28380 gagaccgtct acctcaccaa ccagactgag gtaaaactca cctgcagacc acacaagacc 28440 tatatcatct ggttcttcga gaacacctca tttgcagtct ccaacactca ctgcaacgac 28500 ggtgttgaac ttcccaacaa cctttccagt ggactgagtt acgatacaca tagagctaag 28560 ctcgtcctct acaatccttt tgtagaggga acctaccagt gccagagcgg accttgtact 28620 cacaccttcc atttggtgaa cgtcaccagc agcagcaaca gctcagaaac taaccttcct 28680 tctgatacta acaaacctcg tttcggaggt gagctaaggc ttcccccttc tgaggagggg 28740 gttagcccat acgaagtggt cgggtatttg attttagggg tggtcctggg tgggtgcata 28800 gcggtgctag tcgagatgga cggccaggcc tccgagcagc gcatcctgca actgcgcgtc 28860 cgtcagcagc aggagcgtgc cgccaaggag ctcctcgatg ccatcaacat ccaccagtgc 28920 aagaagggca tcttctgcct ggtcaaacag gcaaagatca cctacgagct cgtgtccggc 28980 ggcaagcagc atcgcctcgc ctatgagctg ccccagcaga agcagaagtt cacctgcatg 29040 gtgggcgtca accccatagt catcacccag cagtcgggcg agaccagcgg ctgcatccac 29100 tgctcctgcg aaagccccga gtgcatctac tccctgctca agaccctttg cggactccgc 29160 gacctcctcc ccatgaactg atgttgatta aaagcccaaa aaccaatcag ccccttcccc 29220 ctaatcattc aataaagatc acttacttga aatctgaaag tatgtctctg gtgtagttgt 29280 tcagcagcac ctcggtaccc tcctcccagc tctggtactc cagtccccgg cgggcggcga 29340 acttcctcca caccttgaaa gggatgtcaa attcctggtc cacaattttc attgtcttcc 29400 ctctcagatg gcaaagaggc tccgggtgga agatgacttc aaccccgtct acccctatgg 29460 ctacgcgcgg atcagaata tccccttcct cactcccccc ttgtctcct ccgatggatt 29520 caaaaacttc ccccctgggg tcctgtcact taactggct catcaatca ccacacaca 29580 tgggatgtc tcacttagg tgggagggg acttgctgta gagcacaga ctggtaacct 29640 aagcgtaaac cctgatgcac ccttgcaagt tgcaagtgat aagctacagc tgctctggc 29700 tcctccattc gaggtcagag atggaagct tgctttaaag gcaggtaatg gattaaagt 29760 actagataat tccattactg gattgactgg attattgaat acacttgtgg tattaactgg 29820 aagggaata ggaacggagg aattaaaaa tgacgatggt gtaacaaca aaggagtcgg 29880 cttgcgtgta agacttggag atgacggcgg gctgacatttt gataaaaagg gtgatttagt 29940 agcctggaat aaaaaagatg acggcgcac cctgtggaca acccctgaca catctccaaa 30000 ttgcaaatg agtacagaaa aggattctaa acttacgttg acacttacaa agtgtggaag 30060 tcaggttctg ggaatgtat ctttacttgc agttacaggt gatatcatc aaatgactgc 30120 tactacaaag aaggatgtaa aaataatcttt actatttgat gagaatggaa ttctattacc 30180 atcttcgtcc cttagcaaag attattggaa ttacagaagt gatgattcta ttgtatctca 30240 aaaatataat aatgcagttc cattcatgcc aaacctgaca gcttatcca aaccaagcgc 30300 tcaaatgca aaaaactatt caagaactaa atcatagt atgtctact taggtgctct 30360 tacctaccaa cctgtaatta tcactattgc atttaatcag gaactgaaa atggatgtgc 30420 ttattctata acatttacct catggca aaagactt catcaccac agtttgatgt 30480 tacatcttttt accttctcat atcttacc agagaacaaa gataaaatgt 30540 tttgaactga atttatgaat ctttattt ttttacacca gcacgggtag tcagttttccc 30600 accaccagcc cattcacag tgtaacaat tctctcagca cgggtggcct taaacaggtc 30660 acagaaccct agtattcaac ctgccaccctc cctcccaaca cacagagtac acagtccttt 30720 ctccccggct ggccttaaaa agcatcatat catgggtaac agcatattc ttaggtgtta 30780 tattccacac gttttcctgt cgagccaaac gctcatcagt gatatata aactccccgg 30840 gcagctcact taagttcatg tcgctgtcca gctgctgagc cacaggctgc tgtccaactt 30900 gcggttgctt aacgggcggc gaaggagaag tccacgccta catgggggta gagtcataat 30960 cgtgcatcag gatagggcgg tggtgctgca gcagcgcgcg aataaactgc tgccgccgcc 31020 gctccgtcct gcaggaatac aacatggcag tggtctcctc agcgatgatt cgcaccgccc 31080 gcagcataag gcgccttgtc ctccgggcac agcagcgcac cctgatctca cttaaatcag 31140 cacagtaact gcagcacagc accacaatat tgttcaaaat cccacagtgc aaggcgctgt 31200 atccaaagct catggcgggg accacagaac ccacgtggcc atcataccac aagcgcaggt 31260 agattaagtg gcgacccctc ataaacacgc tggacataaa cattacctct tttggcatgt 31320 tgtaattcac cacctcccgg taccatataa acctctgatt aaacatggcg ccatccacca 31380 ccatcctaaa ccagctggcc aaaacctgcc cgccggctat acactgcagg gaaccgggac 31440 tggaacaatg acagtggaga gcccaggact cgtaaccatg gatcatcatg ctcgtcatga 31500 tatcaatgtt ggcacaacac aggcacacgt gcatacactt cctcaggatt acaagctcct 31560 cccgcgttag aaccatatcc caggagaaa cccattcctg aatcagcgta aatcccacac 31620 tgcagggaag acctcgcacg taactcacgt tgtgcattgt caaggtgtta cattcgggca 31680 gcaagcggatg atcctccagt atggtagcgc gggtttctgt ctcaaaagga ggtagacgat 31740 ccctactgta cggagtgcgc cgagacaacc gagatcgtgt tggtcgtagt gtcatgccaa 31800 atggaacgcc ggacgtagtc atatttcctg aaggagctcg actgttcctc ggtggacatt 31860 gaaatggatt ctcttgcgta ccttgtcgta cttctgccag cagaaagtgg ctcgggaaca 31920 gcagatacct ttcctcctgc tgtccttccg ctgctgacgc tcagtcatcc aactgagta 31980 cagccattcc cgcaggttct ccagcagctc ctgtgcatct gatgaaaaa aagtcccgtc 32040 gatgcggatt ccccttaaaa catcagccag gacattgtag gccatccccaa tccagttaat 32100 gcatcctgat ctatcatgaa gaggaggtgg gggaagaact ggaagaacca tttttattcc 32160 aagcggtctc gaaggacgat aaagtgcaag tcacgcaggt gacagcgttc cccgccgctg 32220 tgctggtgga aacagacagc caggtcaaaa cccactctat tttcaaggtg ctcgactgtg gcttcgagca gtggctctac gcgtacatcc agcataaga tcacattaaa ggctggacct ccatcgattt catcaatcat caggttacac tcattcacca tccccaggta attctcattt ttccagcctt ggattatttc tacaaattgt tggtgtaagt ccactccgca catgtggaaa agttcccaca gcgccccctc cactttcata atcaggcaga ccttcatatt agaaacagat cctgctgctc caccacctgc agcgtgttca aaacaacaag attcaatgag gttctgccct ctgccctcag ctcacgtctc agcgtcagct gcaaaaagtc actcaagtcc tcagccacta cagctgacaa ttcagagcca gggctaagcg tgggactggc aagcgtgagt gagtaccacc 32760. caaaaactgc atgctggaat aagctctctt tgtgtcaccg gtgatgcctt ccaataggtg agtgataaag cgaggtagtt tttctttaat catttgagta atagaaaagt cctctaaata agtcactagg accccagga ccacaatgtg gtagctgaca gcgtgtcgct caagcatggt tagtagat gagagtctga aaaacagaaa gcatgcacta aaccagagtt gccagtctca ctgaaggaaa aatcactctc tccagcagca aagtgcccac tgggtggccc tctcggacat 33000 acaaaaatcg atccgtgtgg ttaaagagca gcacagttag ctcctgtctt ctcccagcaa 33060 agatcacatc ggactgggtt agtatgcccc tggaatggta gtcattcaag gccataaatc 33120 tgccttggta gccattagga atcagcacgc tcactctcaa gtgaaccaaa accaccccat 33180 gcggaggaat gtggaaagat tctgggcaaa aaaaggtata tctattgcta gtcccttcct 33240 ggacgggagc aatccctcca gggctatcta tgaaagcata cagagattca gccatagctc 33300 agcccgctta ccagtagaca gagagcacag cagtacaagc gccaacagca gcgactgact 33360 acccactgac ccagctccct atttaaaggc accttacact gacgtaatga ccaaaggtct 33420 aaaaaccccg ccaaaaaaac acacacgccc tgggtgtttt tcgcgaaaac acttccgcgt 33480 tctcacttcc tcgtatcgat ttcgtgactc aacttccggg ttcccacgtt acgtcacttc 33540 tgcccttaca tgtaactcag ccgtagggcg ccatcttgcc cacgtccaaa atggcttcca 33600 tgtccggcca cgcctccgcg gcgaccgtta gccgtgcgtc gtgacgtcat ttgcatcacc 33660 gtttctcgtc caatcagcgt tggctccgcc ccaaaaccgt taaaattcaa aagctcattt gcatattaac ttttgtttac tttgtggggt atattattga tgatg <210> 6 <211> 34808 <212> DNA <213> Recombinant simian adenovirus serotype 25 (Recombinant simian adenovirus serotype 25) <400> 6 catcaat aatatacctt attttggatt gaagccaata tgataatgag ggggtggagt ttgtgacgtg gcgcggggcg tgggaacggg gcgggtgacg tagtagtgtg gcggaagtgt gatgttgcaa gtgtggcgga acacatgtaa gcgacggatg tggcaaaagt gacgtttttg gtgtgcgccg gtgtacacag gaagtgacaa ttttcgcgcg gttttaggcg gatgttgtag 240 taaatttggg cgtaaccgag taagattttgg ccattttcgc gggaaaactg aataagagga agtgaaatct gaataatttt gtgttactca tagcgcgtaa tacatggccc gaaaggagcg atgtaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg 480 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg 540 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt 600 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg 660 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg 720 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt 780 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac 840 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg 900 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gagatctggt 960 accgtcgacg cggccgctcg agcctaagct tggtaccatg tttgtgttcc ttgtgttatt 1020 gccactagtc tctagtcagt gtgtgaacct gaccacaaga acccagctgc ctccagccta 1080 caccaacagc tttaccagag gcgtgtacta ccccgacaag gtgttcagat ccagcgtgct 1140 gcactctacc caggacctgt tcctgccttt cttcagcaac gtgacctggt tccacgccat 1200 ccacgtgtcc ggcaccaatg gcaccaagag attcgacaac cccgtgctgc ccttcaacga 1260 cggggtgtac tttgccagca ccgagaagtc caacatcatc agaggctgga tcttcggcac 1320 cacactggac agcaagaccc agagcctgct gatcgtgaac aacgccacca acgtggtcat 1380 caaagtgtgc gagttccagt tctgcaacga ccccttcctg ggcgtctact atcacaagaa 1440 caacaagagc tggatggaaa gcgagttccg ggtgtacagc agcgccaaca actgcacctt 1500 cgagtacgtg tcccagcctt tcctgatgga cctggaaggc aagcagggca acttcaagaa 1560 cctgcgcgag ttcgtgttca agaacatcga cggctacttc aagatctaca gcaagcacac 1620 ccctatcaac ctcgtgcggg atctgcctca gggcttctct gctctggaac ccctggtgga 1680 tctgcccatc ggcatcaaca tcacccggtt tcagacactg ctggccctgc acagaagcta 1740 cctgacacct ggcgatagca gcagcggatg gacagctggt gccgccgctt actatgtggg 1800 ctacctgcag cctagaacct tcctgctgaa gtacaacgag aacggcacca tcaccgacgc 1860 cgtggattgt gctctggatc ctctgagcga gacaaagtgc accctgaagt ccttcaccgt 1920 ggaaaagggc atctaccaga ccagcaactt ccgggtgcag cccaccgaat ccatcgtgcg 1980 gttccccaat atcaccaatc tgtgcccctt cggcgaggtg ttcaatgcca ccagattcgc 2040 ctctgtgtac gcctggaacc ggaagcggat cagcaattgc gtggccgact actccgtgct 2100 gtacaactcc gccagcttca gcaccttcaa gtgctacggc gtgtccccta ccaagctgaa 2160 cgacctgtgc ttcacaaacg tgtacgccga cagcttcgtg atccggggag atgaagtgcg 2220 gcagattgcc cctggacaga caggcaagat cgccgactac aactacaagc tgcccgacga 2280 cttcaccggc tgtgtgattg cctggaacag caacaacctg gactccaaag tcggcggcaa 2340 ctacaattac ctgtaccggc tgttccggaa gtccaatctg aagcccttcg agcgggacat 2400 ctccaccgag atctatcagg ccggcagcac cccttgtaac ggcgtggaag gcttcaactg 2460 ctacttccca ctgcagtcct acggctttca gcccacaaat ggcgtgggct atcagcccta 2520 cagagtggtg gtgctgagct tcgaactgct gcatgcccct gccacagtgt gcggccctaa 2580 gaaaagcacc aatctcgtga agaacaaatg cgtgaacttc aacttcaacg gcctgaccgg 2640 caccggcgtg ctgacagaga gcaacaagaa gttcctgcca ttccagcagt ttggccggga 2700 tattgccgat accacagacg ccgtacgaga tccccagaca ctggaaatcc tggacatcac 2760 cccttgcagc ttcggcggag tgtctgtgat cacccctggc accaacacca gcaatcaggt 2820 ggcagtgctg taccaggacg tgaactgtac cgaagtgccc gtggccattc acgccgatca 2880 gctgacacct acatggcggg tgtactccac cggcagcaat gtgtttcaga ccagagccgg 2940 ctgtctgatc ggagccgagc acgtgaacaa tagctacgag tgcgacatcc ccatcggcgc 3000 tggcatctgt gccagctacc agacacagac aaacagcccc agacgggcca gatctgtggc 3060 cagccagagc atcattgcct acacaatgtc tctgggcgcc gagaacagcg tggcctactc 3120 caacaactct atcgctatcc ccaccaactt caccatcagc gtgaccacag agatcctgcc 3180 tgtgtccatg accaagacca gcgtggactg caccatgtac atctgcggcg attccaccga 3240 gtgctccaac ctgctgctgc agtacggcag cttctgcacc cagctgaata gagccctgac 3300 agggatcgcc gtggaacagg acaagaacac ccaagaggtg ttcgcccaag tgaagcagat ctacaagacc cctcctatca aggacttcgg cggcttcaat ttcagccaga ttctgcccga tcctagcaag cccagcaagc ggagcttcat cgaggacctg ctgttcaaca aagtgacact ggccgacgcc ggcttcatca agcagtatgg cgattgtctg ggcgacattg ccgccaggga 3540 tctgatttgc gcccagaagt ttaacggact gacagtgctg ccaccactgc tgaccgatga gatgatcgcc cagtacacat ctgccctgct ggccggcaca atcacaagcg gctggacatt tggagctggc gccgctctgc agatcccctt tgctatgcag atggcctacc ggttcaacgg 3720 catcggagtg acccagaatg tgctgtacga gaaccagaag ctgatcgcca accagttcaa cagcgccatc ggcaagatcc aggacagcct cagcagcaca gcaagcgccc tgggaagct gcaggacgtg gtcaaccaga atgcccaggc actgaacacc ctggtcaagc agctgtcctc caacttcggc gccatcagct ctgtgctgaa cgacatcctg agcagactgg acaaggtgga agccgaggtg cagatcgaca cactgatcac cggaaggctg cagtccctgc agcctacgt tacccagcag ctgatcagag ccgccgagat tagagcctct gccaatctgg ccgccaccaa 4080 gatgtctgag tgtgtgctgg gccagagcaa gagagtggac ttttgcggca agggctacca 4140 cctgatgagc ttccctcagt ctgcccctca cggcgtggtg tttctgcacg tgacatacgt 4200 gcccgctcaa gagaagaatt tcaccaccgc tccagccatc tgccacgacg gcaaagccca 4260 ctttcctaga gaaggcgtgt tcgtgtccaa cggcacccat tggttcgtga cccagcggaa 4320 cttctacgag ccccagatca tcaccaccga caacaccttc gtgtctggca actgcgacgt 4380 cgtgatcggc attgtgaaca ataccgtgta cgaccctctg cagcccgagc tggacagctt 4440 caaagaggaa ctggataagt actttaagaa ccacacaagc cccgacgtgg acctgggcga 4500 catcagcgga atcaatgcca gcgtcgtgaa catccagaaa gagatcgacc ggctgaacga 4560 ggtggccaag aatctgaacg agagcctgat cgacctgcaa gaactgggga agtacgagca 4620 gtacatcaag tggccctggt acatctggct gggctttatc gccggactga ttgccatcgt 4680 gatggtcaca atcatgctgt gttgcatgac cagctgctgt agctgcctga agggctgttg 4740 tagctgtggc agctgctgca agttcgacga ggacgattct gagcccgtgc tcaaaggagt 4800 4860. 4860. 4860. 4860. 4860. 4860. 4860. 4860. 4860. 4860 ccataccaca tttgtagagg ttttacttgc tttaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaata agcaatagca tcacaaattt cacaataa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc ggatctgggc gtggttaagg gtgggaaaga fathergg tgggggtctt atgtagtttt gtatctgttt tgcagcagcc gccgccgcca tgagcaccaa ctcgtttgat ggaagcattg tgagctcata tttgacaacg cgcatgcccc catgggccgg ggtgcgtcag aatgtgatgg gctccagcat tgatggtcgc 5280 ccgtcctgc ccgcaaactc tactaccttg acctacgaga ccgtgtctgg aacgccgttg 5340 gagactgcag cctccgccgc cgcttcagcc gctgcagcca ccgcccgcgg gattgtgact 5400 gactttgctt tcctgagccc gcttgcaagc agtgcagctt cccgttcatc cgcccgcgat 5460 gacaagttga cggctctttt ggcacaattg gattctttga cccgggaact taatgtcgtt 5520 tctcagcagc tgttggatct gcgccagcag gtttctgccc tgaaggcttc ctcccctccc 5580 aatgcggttt aaaacataaa taaaaaacca gactctgttt ggatttggat caagcaagtg 5640 tcttgctgtc tttatttagg ggttttgcgc gcgcggtagg cccgggacca gcggtctcgg 5700 tcgttgaggg tcctgtgtat tttttccagg acgtggtaaa ggtgactctg gatgttcaga 5760 tacatgggca taagcccgtc tctggggtgg aggtagcacc actgcagagc ttcatgctgc 5820 ggggtggtgt tgtagatgat ccagtcgtag caggagcgct gggcgtggtg cctaaaaatg 5880 tctttcagta gcaagctgat tgccaggggc aggcccttgg tgtaagtgtt tacaaagcgg 5940 ttaagctggg atgggtgcat acgtggggat atgagatgca tcttggactg tatttttagg 6000 ttggctatgt tcccagccat atccctccgg ggattcatgt tgtgcagaac caccagcaca 6060 gtgtatccgg tgcacttggg aaatttgtca tgtagcttag aaggaaatgc gtggaagaac 6120 ttggagacgc ccttgtgacc tccaagattt tccatgcatt cgtccataat gatggcaatg 6180 ggcccacggg cggcggcctg ggcgaagata tttctgggat cactaacgtc atagttgtgt 6240 tccaggatga gatcgtcata ggccattttt acaaagcgcg ggcggagggt gccagactgc 6300 ggtataatgg ttccatccgg cccaggggcg tagttaccct cacagatttg catttcccac 6360 gctttgagtt cagatggggg gatcatgtct acctgcgggg cgatgaagaa aacggtttcc 6420 ggggtagggg agatcagctg ggaagaaagc aggttcctga gcagctgcga cttaccgcag 6480 ccggtgggcc cgtaaatcac acctattacc ggctgcaact ggtagttaag agagctgcag 6540 ctgccgtcat ccctgagcag gggggccact tcgttaagca tgtccctgac tcgcatgttt 6600 tccctgacca aatccgccag aaggcgctcg ccgcccagcg atagcagttc ttgcaaggaa 6660 gcaaagtttt tcaacggttt gagaccgtcc gccgtaggca tgcttttgag cgtttgacca 6720 agcagttcca ggcggtccca cagctcggtc acctgctcta cggcatctcg atccagcata 6780 tctcctcgtt tcgcgggttg gggcggcttt cgctgtacgg cagtagtcgg tgctcgtcca 6840 gacgggccag ggtcatgtct ttccacgggc gcagggtcct cgtcagcgta gtctgggtca 6900 cggtgaaggg gtgcgctccg ggctgcgcgc tggccagggt gcgcttgagg ctggtcctgc 6960 tggtgctgaa gcgctgccgg tcttcgccct gcgcgtcggc caggtagcat ttgaccatgg 7020 tgtcatagtc cagcccctcc gcggcgtggc ccttggcgcg cagcttgccc ttggaggagg 7080 cgccgcacga ggggcagtgc agacttttga gggcgtagag cttgggcgcg agaaataccg 7140 attccgggga gtaggcatcc gcgccgcagg ccccgcagac ggtctcgcat tccacgagcc 7200 aggtgagctc tggccgttcg gggtcaaaaa ccaggtttcc cccatgcttt ttgatgcgtt 7260 tcttacctct ggtttccatg agccggtgtc cacgctcggt gacgaaaagg ctgtccgtgt 7320 ccccgtatac agacttgaga ggcctgtcct cgagcggtgt tccgcggtcc tcctcgtata 7380 gaaactcgga ccactctgag acaaaggctc gcgtccaggc cagcacgaag gaggctaagt 7440 gggaggggta gcggtcgttg tccactaggg ggtccactcg ctccagggtg tgaagacaca 7500 tgtcgccctc ttcggcatca aggaaggtga ttggtttgta ggtgtaggcc acgtgaccgg 7560 gtgttcctga aggggggcta taaaaggggg tgggggcgcg ttcgtcctca ctctcttccg 7620 catcgctgtc tgcgagggcc agctgttggg gtgagtactc cctctgaaaa gcgggcatga 7680 cttctgcgct aagattgtca gtttccaaaa acgaggagga tttgatattc acctggcccg 7740 cggtgatgcc tttgagggtg gccgcatcca tctggtcaga aaagacaatc tttttgttgt 7800 caagcttggt ggcaaacgac ccgtagaggg cgttggacag caacttggcg atggagcgca 7860 gggtttggtt tttgtcgcga tcggcgcgct ccttggccgc gatgtttagc tgcacgtatt 7920 cgcgcgcaac gcaccgccat tcgggaaaga cggtggtgcg ctcgtcgggc accaggtgca 7980 cgcgccaacc gcggttgtgc agggtgacaa ggtcaacgct ggtggctacc tctccgcgta 8040 ggcgctcgtt ggtccagcag aggcggccgc ccttgcgcga gcagaatggc ggtagggggt 8100 ctagctgcgt ctcgtccggg gggtctgcgt ccacggtaaa gaccccgggc agcaggcgcg 8160 cgtcgaagta gtctatcttg catccttgca agtctagcgc ctgctgccat gcgcgggcgg 8220 caagcgcgcg ctcgtatggg ttgagtgggg gaccccatgg catggggtgg gtgagcgcgg 8280 aggcgtacat gccgcaaatg tcgtaaacgt agaggggctc tctgagtatt ccaagatatg 8340 tagggtagca tcttccaccg cggatgctgg cgcgcacgta atcgtatagt tcgtgcgagg 8400 gagcgaggag gtcgggaccg aggttgctac gggcgggctg ctctgctcgg aagactatct 8460 gcctgaagat ggcatgtgag ttggatgata tggttggacg ctggaagacg ttgaagctgg 8520 cgtctgtgag acctaccgcg tcacgcacga aggaggcgta ggagtcgcgc agcttgttga 8580 ccagctcggc ggtgacctgc acgtctaggg cgcagtagtc cagggtttcc ttgatgatgt 8640 catacttatc ctgtcccttt tttttccaca gctcgcggtt gaggacaaac tcttcgcggt ctttccagta ctcttggatc ggaaacccgt cggcctccga acggtaagag cctagcatgt 8760. agaactggtt gacggcctgg taggcgcagc atcccttttc tacgggtagc gcgtatgcct 8820 gcgcggcctt ccggagcgag gtgtgggtga gcgcaaaggt gtccctgacc atgactttga 8880 ggtactggta tttgaagtca gtgtcgtcgc atccgccctg ctcccagagc aaaaagtccg 8940. tgcgcttttt ggacgcgga tttggcaggg cgaaggtgac atcgttgaag agtatctttc ccgcgcgagg cataaagttg cgtgtgatgc ggaagggtcc cggcacctcg gaacggttgt 9060 taattacctg ggcggcgagc acgatctcgt caaagccgtt gatgttgtg cccacaatgt 9120 aaagttccaa gaagcgcggg atgcccttga tggaggcaa tttttaagt tcctcgtagg 9180 tgagctctc agggagctg agcccgtgct ctgaagggc ccagtctgca agatgaggggt 9240 tggaagcgac gatgagctc cacaggtcac gggccattag cattgcagg tggtcgcgaa 9300 aggtcctaaa ctggcgacct atggccatttt ttctggggt gatgcagtag aaggtaagcg 9360 ggtcttgttc ccagcggtcc catccaggt tcgcggctag gtctcgcgcg gcagtcacta 9420 gaggctcatc tccgccgaac ttcatgacca gcatgaaggg cacgagctgc ttcccaagg 9480 cccccatcca agtataggtc tctacatcgt aggtgacaaa gagacgctcg gtgcgaggat 9540 gcgagccgat cgggaagaac tggatctccc gccaccaatt ggaggagtgg ctattgatgt 9600 ggtgaaagta gaagtccctg cgacggggccg aacactcgtg ctggctttg taaaaacgtg 9660 cgcagtactg gcagcggtgc acgggctgta catcctgcac gaggttgacc tgacgaccgc 9720 gcacaaggaa gcagagtggg aatttgagcc cctcgcctgg cgggttttggc tggtggtctt 9780 ctacttcggc tgcttgtcct tgaccgtctg gctgctcgag gggagttacg gtggatcgga 9840 ccaccacgcc gcgcgagccc aaagtccaga tgtccgcgcg cggcggtcgg agcttgatga 9900 caacatcgcg cagatgggag ctgtccatgg tctggagctc ccgcggcgtc aggtcaggcg 9960 ggagctcctg caggtttacc tcgcatagac gggtcagggc gcgggctaga tccaggtgat 10020 acctaatttc caggggctgg ttggtggcgg cgtcgatggc ttgcaagagg ccgcatcccc 10080 gcggcgcgac tacggtaccg cgcggcgggc ggtgggccgc gggggtgtcc ttggatgatg 10140 catctaaaag cggtgacgcg ggcgagcccc cggaggtagg gggggctccg gacccgccgg 10200 gagagggggc aggggcacgt cggcgccgcg cgcgggcagg agctggtgct gcgcgcgtag 10260 gttgctggcg aacgcgacga cgcggcggtt gatctcctga atctggcgcc tctgcgtgaa 10320 gacgacgggc ccggtgagct tgaacctgaa agagagttcg acagaatcaa tttcggtgtc 10380 gttgacggcg gcctggcgca aaatctcctg cacgtctcct gagttgtctt gataggcgat 10440 ctcggccatg aactgctcga tctcttcctc ctggagatct ccgcgtccgg ctcgctccac 10500 ggtggcggcg aggtcgttgg aaatgcgggc catgagctgc gagaaggcgt tgaggcctcc 10560 ctcgttccag acgcggctgt agaccacgcc cccttcggca tcgcgggcgc gcatgaccac 10620 ctgcgcgaga ttgagctcca cgtgccgggc gaagacggcg tagtttcgca ggcgctgaaa 10680 gaggtagttg agggtggtgg cggtgtgttc tgccacgaag aagtacataa cccagcgtcg 10740 caacgtggat tcgttgatat cccccaaggc ctcaaggcgc tccatggcct cgtagaagtc 10800 cacggcgaag ttgaaaaact gggagttgcg cgccgacacg gttaactcct cctccagaag 10860 acggatgagc tcggcgacag tgtcgcgcac ctcgcgctca aaggctacag gggcctcttc 10920 ttcttcttca atctcctctt ccataagggc ctccccttct tcttcttctg gcggcggtgg 10980 gggagggggg acacggcggc gacgacggcg caccgggagg cggtcgacaa agcgctcgat 11040 catctccccg cggcgacggc gcatggtctc ggtgacggcg cggccgttct cgcgggggcg 11100 cagttggaag acgccgcccg tcatgtcccg gttatgggtt ggcggggggc tgccatgcgg 11160 cagggatacg gcgctaacga tgcatctcaa caattgttgt gtaggtactc cgccgccgag 11220 ggacctgagc gagtccgcat cgaccggatc ggaaaacctc tcgagaaagg cgtctaacca 11280 gtcacagtcg caaggtaggc tgagcaccgt ggcgggcggc agcgggcggc ggtcggggtt 11340 gtttctggcg gaggtgctgc tgatgatgta attaaagtag gcggtcttga gacggcggat 11400 ggtcgacaga agcaccatgt ccttgggtcc ggcctgctga atgcgcaggc ggtcggccat 11460 gccccaggct tcgttttgac atcggcgcag gtctttgtag tagtcttgca tgagcctttc 11520 taccggcact tcttcttctc cttcctcttg tcctgcatct cttgcatcta tcgctgcggc 11580 ggcggcggag tttggccgta ggtggcgccc tcttcctccc atgcgtgtga ccccgaagcc 11640 cctcatcggc tgaagcaggg ctaggtcggc gacaacgcgc tcggctaata tggcctgctg 11700 cacctgcgtg agggtagact ggaagtcatc catgtccaca aagcggtggt atgcgcccgt 11760 gttgatggtg taagtgcagt tggccataac ggaccagtta acggtctggt gacccggctg 11820 cgagagctcg gtgtacctga gacgcgagta agccctcgag tcaaatacgt agtcgttgca 11880 agtccgcacc aggtactggt atcccaccaa aaagtgcggc ggcggctggc ggtagagggg 11940 ccagcgtagg gtggccgggg ctccggggggc gagatcttcc aacataaggc gatgatatcc gtagatgtac ctggacatcc aggtgatgcc ggcggcggtg gtggaggcgc gcggaagtc 12060 gcggacgcgg ttccagatgt tgcgcagcgg caaaaagtgc tccatggtcg ggacgctctg 12120 gccggtcagg cgcgcgcaat cgttgacgct ctagaccgtg caaaggaga gcctgtaagc 12180 gggcactctt ccgtggtctg gtggataaat tcgcaagggt atcatggcgg acgaccgggg 12240 ttcgagcccc gtatccggcc gtccgccgtg atccatgcgg ttaccgccccg cgtgtcgaac 12300. ccaggtgtgc gacgtcagac aacgggggag tgctcctttt ggcttccttc caggcgcggc 12360 ggctgctgcg ctagcttttt tggccactgg ccgcgcgcag cgtagcggt tagctgga 12420 agcgaaagca ttaagtggct cgctccctgt agccggaggg ttattttcca agggttgagt cggcgggaccc ccggttcgag tctcggaccg gccggactgc ggcgaacggg ggtttgcctc 12540. cccgtcatgc aagaccccgc ttgcaaattc ctccggaaac agggacgagc cccttttttg 12600 cttttcccag atgcatccgg tgctgcggca gatgcgcccc cctcctcagc agcggcaaga 12660 gcaagagcag cggcagacat gcagggcacc ctcccctcct cctaccgcgt caggaggggc 12720 gacatccgcg gttgacgcgg cagcagatgg tgattacgaa cccccgcggc gccgggcccg 12780 gcactacctg gacttggagg agggcgaggg cctggcgcgg ctaggagcgc cctctcctga 12840 gcggcaccca agggtgcagc tgaagcgtga tacgcgtgag gcgtacgtgc cgcggcagaa 12900 cctgtttcgc gaccgcgagg gagaggagcc cgaggagatg cgggatcgaa agttccacgc 12960 agggcgcgag ctgcggcatg gcctgaatcg cgagcggttg ctgcgcgagg aggactttga 13020 gcccgacgcg cgaaccggga ttagtcccgc gcgcgcacac gtggcggccg ccgacctggt 13080 aaccgcatac gagcagacgg tgaaccagga gattaacttt caaaaaagct ttaacaacca 13140 cgtgcgtacg cttgtggcgc gcgaggaggt ggctatagga ctgatgcatc tgtgggactt 13200 tgtaagcgcg ctggagcaaa acccaaatag caagccgctc atggcgcagc tgttccttat 13260 agtgcagcac aggravate acgaggcatt cagggatgcg ctgctaaca tagtagcc cgagggccgc tggctgctcg atttgataaa catcctgcag agcatagtgg tgcaggagcg 13440. cagcttgagc ctggctgaca aggtggccgc catcaactat tccatgctta gcctgggcaa gttttacgcc cgcaagatat accatacccc ttacgttccc atagcaagg aggtaagat cgaggggttc tacatgcgca tggcgctgaa ggtgcttacc ttgagcgacg acctgggcgt ttatcgcaac gagcgcatcc acaaggccgt gagcgtgagc cggcggcgcg agctcagcga ccgcgagctg atgcacagcc tgcaaagggc cctggctggc acgggcagcg gcgatagaga 13680 ggccgagtcc tactttgacg cggggcgctga cctgcgctgg gccccaagcc gacgcgccct ggaggcagct ggggccggac ctgggctggc ggtggcaccc gcgcgcgctg gcaacgtcgg 13800 cggcgtggag gaatatgacg aggacgatga gtacgagcca gaggacggcg agtactagc ggtgatgttt ctgatcagat gatgcaagac gcaacggacc cggcggtgcg ggcggcgctg 13920 cagagccagc cgtccggcct taactccacg gacgactggc gccaggtcat ggaccgcatc 13980. atgtcgctga ctgcgcgcaa tcctgacgcg ttccggcagc agccgcaggc caaccggctc 14040. tccgcaattc tggaagcggt ggtcccggcg cgcgcaaacc ccacgcacga gaaggtgctg gcgatcgtaa acgcgctggc cgaaaacagg gccatccggc ccgacgaggc cggcctggtc 14160 tacgacgcgc tgcttcagcg cgtggctcgt tacaacagcg gcaacgtgca gaccaacctg gaccggctgg tgggggatgt gcgcgaggcc gtggcgcagc gtgagcgcgc gcagcagcag 14280 14340. ggcaacctgg gctccatggt tgcactaaac gccttcctga gtacacagcc cgccaacgtg ccgcggggac aggaggacta caccaacttt gtgagcgcac tgcggctaat ggtgactgag 14400 acaccgcaaa gtgaggtgta ccagtctggg ccagactatt ttttccagac cagtagacaa ggcctgcaga ccgtaaacct gagccaggct ttcaaaaact tgcaggggct gtggggggtg cgggctccca caggcgaccg cgcgaccgtg tctagcttgc tgacgcccaa ctcgcgcctg ttgctgctgc fathercgcc cttcacggac agtggcagcg tgtcccggga cacataccta ggtcacttgc tgacactgta ccgcgaggcc ataggtcagg cgcatgtgga cgagcatact 14700 ttccaggaga ttacaagtgt cagccgcgcg ctggggcagg aggacacggg cagcctggag 14760 gcaaccctaa actacctgct gaccaaccgg cggcagaaga tcccctcgtt gcacagttta 14820 aacagcgagg aggagcgcat tttgcgctac gtgcagcaga gcgtgagcct taacctgatg 14880 cgcgacgggg taacgcccag cgtggcgctg gacatgaccg cgcgcaacat ggaaccgggc 14940 atgtatgcct caaaccggcc gtttatcaac cgcctaatgg actacttgca tcgcgcggcc 15000 gccgtgaacc ccgagtattt caccaatgcc atcttgaacc cgcactggct accgccccct 15060 ggtttctaca ccgggggatt cgaggtgccc gagggtaacg atggattcct ctgggacgac 15120 atagacgaca gcgtgttttc cccgcaaccg cagaccctgc tagagttgca acagcgcgag 15180 caggcagagg cggcgctgcg aaaggaaagc ttccgcaggc caagcagctt gtccgatcta 15240 ggcgctgcgg ccccgcggtc agatgctagt agcccatttc caagcttgat agggtctctt 15300 accagcactc gcaccacccg cccgcgcctg ctgggcgagg aggagtacct aaacaactcg 15360 ctgctgcagc cgcagcgcga aaaaacctg cctccggcat ttcccaacaa cgggatagag 15420 agcctagtgg acaagatgag tagatggaag acgtacgcgc aggagcacag ggacgtgcca 15480 ggcccgcgcc cgcccacccg tcgtcaaagg cacgaccgtc agcggggtct ggtgtgggag 15540 gacgatgact cggcagacga cagcagcgtc ctggatttgg gagggagtgg caacccgttt 15600 gcgcaccttc gcccccaggct ggggagaatg ttttaaaaaa aaaaaaagca tgatgcaaaa 15660 taaaaaactc accaaggcca tggcaccgag cgttggtttt cttgtattcc ccttagtatg 15720 cggcgcgcgg cgatgtatga ggaaggtcct cctccctcct acgagagtgt ggtgagcgcg 15780 gcgccagtgg cggcggcgct gggttctccc ttcgatgctc ccctggaccc gccgtttgtg 15840 cctccgcggt acctgcggcc taccgggggg agaaacagca tccgttactc tgagttggca 15900 cccctattcg acaccacccg tgtgtacctg gtggacaaca agtcaacgga tgtggcatcc 15960 ctgaactacc agaacgacca cagcaacttt ctgaccacgg tcattcaaaa caatgactac 16020 agcccgggg aggcaagcac acagaccactc aatcttgacg accggtcgca ctggggcggc 16080 gacctgaaaa ccatcctgca taccaacatg ccaaatgtga acgagttcat gtttaccaat 16140 aagtttaagg cgcgggtgat ggtgtcgcgc ttgcctacta aggacaatca ggtggagctg 16200 aaatacgagt gggtggagtt cacgctgccc gagggcaact actccgagac catgaccata 16260 gaccttatga aaacgcgat cgtggagcac tacttgaaag tgggcagaca gaacggggtt 16320 ctggaaagcg acatcggggt aaagtttgac acccgcaact tcagactggg gtttgacccc 16380 gtcactggtc ttgtcatgcc tggggtatat aaaacgaag ccttccatcc agacatcatt 16440 ttgctgccag gatgcggggt ggacttcacc cacagccgcc tgagcaactt gttgggcatc 16500 cgcaagcggc aacccttcca ggagggcttt aggatcacct acgatgatct ggagggtggt 16560 16620 cagggcgggg gtggcgcagg cggcagcaac agcagtggca gcggcgcgga agagaactcc 16680 aacgcggcag ccgcggcaat gcagccggtg gaggacatga acgatcatgc cattcgcggc 16740 gacacctttg ccacacgggc tgaggagaag cgcgctgagg ccgaagcagc ggccgaagct 16800 gccgcccccg ctgcgcaacc cgaggtcgag aagcctcaga agaaaccggt gatcaaaccc 16860 ctgacagagg acagcaagaa acgcagttac aacctaataa gcaatgacag caccttcacc 16920 cagtaccgca gctggtacct tgcatacaac tacggcgacc ctcagaccgg aatccgctca 16980 tggaccctgc tttgcactcc tgacgtaacc tgcggctcgg agcaggtcta ctggtcgttg 17040 ccagacatga tgcaagaccc cgtgaccttc cgctccacgc gccagatcag caactttccg 17100 gtggtgggcg ccgagctgtt gcccgtgcac tccaagagct tctacaacga ccaggccgtc 17160 tactcccaac tcatccgcca gtttacctct ctgacccacg tgttcaatcg ctttcccgag 17220 aaccagattt tggcgcgccc gccagccccc accatcacca ccgtcagtga aaacgttcct 17280 gctctcacag atcacgggac gctaccgctg cgcaacagca tcggaggagt ccagcgagtg 17340 accattactg acgccagacg ccgcacctgc ccctacgttt acaaggccct gggcatagtc 17400 tcgccgcgcg tcctatcgag ccgcactttt tgagcaagca tgtccatcct tatatcgccc 17460 agcaataaca caggctgggg cctgcgcttc ccaagcaaga tgtttggcgg ggccaagaag 17520 cgctccgacc aacacccagt gcgcgtgcgc gggcactacc gcgcgccctg gggcgcgcac 17580 aaacgcggcc gcactgggcg caccaccgtc gatgacgcca tcgacgcggt ggtggaggag 17640 gcgcgcaact acacgcccac gccgccacca gtgtccacag tggacgcggc cattcagacc 17700 gtggtgcgcg gagcccggcg ctatgctaaa atgaagagac ggcggaggcg cgtagcacgt 17760 cgccaccgcc gccgacccgg cactgccgcc caacgcgcgg cggcggccct gcttaaccgc 17820 gcacgtcgca ccggccgacg ggcggccatg cgggccgctc gaaggctggc cgcgggtatt 17880 gtcactgtgc cccccaggtc caggcgacga gcggccgccg cagcagccgc ggccattagt 17940 gctatgactc agggtcgcag gggcaacgtg tattgggtgc gcgactcggt tagcggcctg 18000 cgcgtgcccg tgcgcacccg ccccccgcgc aactagattg caagaaaaaa ctacttagac 18060 tcgtactgtt gtatgtatcc agcggcggcg gcgcgcaacg aagctatgtc caagcgcaaa 18120 atcaaagaag agatgctcca ggtcatcgcg ccggagatct atggcccccc gaaaggaa 18180 gagcaggatt acaagccccg aaagctaaag cgggtcaaaa agaaaaagaa agatgatgatgat 18240 gatgaacttg acgacgaggt ggaactgctg cacgctaccg cgcccaggcg acgggtacag 18300 tggaaaggtc gacgcgtaaa acgtgttttg cgacccggca ccaccgtagt ctttacgccc 18360 ggtgagcgct ccacccgcac ctacaagcgc gtgtatgatg aggtgtacgg cgacgaggac 18420 ctgcttgagc aggccaacga gcgctcggg gagtttgcct acggaaagcg gcataaggac 18480 atgctggcgt tgccgctgga cgagggcaac ccaacaccta gcctaaagcc cgtaacactg 18540 cagcaggtgc tgcccgcgct tgcaccgtcc gaaaaaagc gcggcctaaa gcgcgagtct 18600 ggtgacttgg cacccaccgt gcagctgatg gtacccaagc gccagcgact ggaagatgtc 18660 ttggaaaaaa tgaccgtgga acctgggctg gagcccgagg tccgcgtgcg gccaatcaag 18720 caggtggcgc cgggactggg cgtgcagacc gtggacgttc agatacccac taccagtagc 18780 accagtattg ccaccgccac agagggcatg gagacacaaa cgtccccggt tgcctcagcg 18840 gtggcggatg ccgcggtgca ggcggtcgct gcggccgcgt ccaagacctc tacggaggtg 18900 caaacggacc cgtggatgtt tcgcgtttca gccccccggc gcccgcgccg ttcgaggaag 18960 tacggcgccg ccagcgcgct actgcccgaa tatgccctac atccttccat tgcgcctacc 19020 cccggctatc gtggctacac ctaccgcccc agaagacgag caactacccg acgccgaacc 19080 accactggaa cccgccgccg ccgtcgccgt cgccagcccg tgctggcccc gatttccgtg 19140 cgcagggtgg ctcgcgaagg aggcaggacc ctggtgctgc caacagcgcg ctaccacccc 19200 agcatcgttt aaaagccggt ctttgtggtt cttgcagata tggccctcac ctgccgcctc 19260 cgtttcccgg tgccgggatt ccgaggaaga atgcaccgta ggaggggcat ggccggccac 19320 ggcctgacgg gcggcatgcg tcgtgcgcac caccggcggc ggcgcgcgtc gcaccgtcgc 19380 atgcgcggcg gtatcctgcc cctccttatt ccactgatcg ccgcggcgat tggcgccgtg 19440 cccggaattg catccgtggc cttgcaggcg cagagacact gattaaaaac aagttgcatg 19500 tggaaaaatc aaaataaaaa gtctggactc tcacgctcgc ttggtcctgt aactattttg 19560 tagaatggaa gacatcaact ttgcgtctct ggccccgcga cacggctcgc gcccgttcat 19620 gggaaactgg caagatatcg gcaccagcaa tatgagcggt ggcgccttca gctggggctc 19680 gctgtggagc ggcattaaaa atttcggttc caccgttaag aactatggca gcaaggcctg 19740 gaacagcagc acaggccaga tgctgaggga taagttgaaa gagcaaaatt tccaacaaaa 19800 ggtggtagat ggcctggcct ctggcattag cggggtggtg gacctggcca accaggcagt 19860 gcaaaataag attaacagta agcttgatcc ccgccctccc gtagaggagc ctccaccggc 19920 cgtggagaca gtgtctccag aggggcgtgg cgaaaagcgt ccgcgccccg acagggaaga 19980 aactctggtg acgcaaatag acgagcctcc ctcgtacgag gaggcactaa agcaaggcct 20040 gcccaccacc cgtcccatcg cgcccatggc taccggagtg ctgggccagc acacacccgt 20100 aacgctggac ctgcctcccc ccgccgacac ccagcagaaa cctgtgctgc caggcccgac 20160 cgccgttgtt gtaacccgtc ctagccgcgc gtccctgcgc cgcgccgcca gcggtccgcg 20220 atcgttgcgg cccgtagcca gtggcaactg gcaaagcaca ctgaacagca tcgtgggtct 20280 gggggtgcaa tccctgaagc gccgacgatg cttctgatag ctaacgtgtc gtatgtgtgt 20340 catgtatgcg tccatgtcgc cgccagagga gctgctgagc cgccgcgcgc ccgctttcca 20400 agatggctac cccttcgatg atgccgcagt ggtcttacat gcacatctcg ggccaggacg 20460 cctcggagta cctgagcccc gggctggtgc agtttgcccg cgccaccgag acgtacttca 20520 gcctgaataa caagtttaga aaccccacgg tggcgcctac gcacgacgtg accacagacc 20580 ggtcccagcg tttgacgctg cggttcatcc ctgtggaccg tgaggatact gcgtactcgt 20640 acaaggcgcg gttcacccta gctgtgggtg ataaccgtgt gctggacatg gcttccacgt 20700 actttgacat ccgcggcgtg ctggacaggg gccctacttt taagccctac tctggcactg 20760 cctacaacgc cctggctccc aagggtgccc caaatccttg cgaatgggat gaagctgcta 20820 ctgctcttga aataaccta gaagagagg acgatgacaa cgaagcaga gtagacgagc 20880 aagctgagca gcaaaaaact cacgtatttg ggcaggcgcc ttatctggt ataaatatta 20940 caaagggg tattcaaata ggtgtcgaag gtcaaacacc taaatatgcc gataaaacat 21000 ttcaacctga acctcaaata ggagaatctc agtggtacga aacagaaatt aatcatgcag 21060 ctgggagagt cctaaaaaag actaccccaa tgaaaccatg ttacggttca tatgcaaaac 21120 ccacaaatga aaatggaggg caaggcattc ttgtaaagca acaaaatgga aagctagaaa 21180 gtcaagtgga aatgcaattt ttctcaacta ctgaggcagc cgcaggcaat ggtgataact 21240 tgactcctaa agtggtattg tacagtgaag atgtagatat agaaacccca gacactcata 21300 tttcttacat gcccactatt aaaggta actcacgaga actaatgggc caacaatcta 21360 tgcccaacag gcctaattac attgctttta gggacaattt tattggtcta atgtattaca 21420 acagcacggg taatatgggt gttctggcgg gccaagcatc gcagttgaat gctgttgtag 21480 atttgcaaga cagaaaca gagctttcat accagctttt gcttgattcc attggtgata 21540 gaaccaggta cttttctatg tggaatcagg ctgttgacag ctatgatcca gatgttagaa 21600 ttattgaaaa tcatggaact gaagatgaac ttccaaatta ctgctttcca ctgggaggtg 21660 tgattaatac agagactctt accaaggtaa aacctaaaac aggtcaggaa aatggatggg 21720 aaaaagatgc tacagaattt tcagataaaa atgaaataag agttggaaat aattttgcca 21780 tggaaatcaa tctaaatgcc aacctgtgga gaaatttcct gtactccaac atagcgctgt 21840 atttgcccga caagctaaag tacagtcctt ccaacgtaaa aatttctgat aacccaaaca 21900 cctacgacta catgaacaag cgagtggtgg ctcccgggct agtggactgc tacattaacc 21960 ttggagcacg ctggtccctt gactatatgg acaacgtcaa cccatttaac caccaccgca 22020 atgctggcct gcgctaccgc tcaatgttgc tgggcaatgg tcgctatgtg cccttccaca 22080 tccaggtgcc tcagaagttc tttgccatta aaaacctcct tctcctgccg ggctcataca 22140 cctacgagtg gaacttcagg aaggatgtta acatggttct gcagagctcc ctaggaaatg 22200 acctaagggt tgacggagcc agcattaagt ttgatagcat ttgcctttac gccaccttct 22260 tccccatggc ccacaacacc gcctccacgc ttgaggccat gcttagaaac gacaccaacg 22320 accagtcctt taacgactat ctctccgccg ccaacatgct ctaccctata cccgccaacg 22380 ctaccaacgt gcccatatcc atcccctccc gcaactgggc ggctttccgc ggctgggcct 22440 tcacgcgcct taagactaag gaaaccccat cactgggctc gggctacgac ccttattaca 22500 cctactctgg ctctataccc tacctagatg gaacctttta cctcaaccac acctttaaga 22560 aggtggccat tacctttgac tcttctgtca gctggcctgg caatgaccgc ctgcttaccc 22620 ccaacgagtt tgaaattaag cgctcagttg acggggaggg ttacaacgtt gcccagtgta 22680 acatgaccaa agactggttc ctggtacaaa tgctagctaa ctataacatt ggctaccagg 22740 gcttctatat cccagagagc tacaaggacc gcatgtactc cttctttaga aacttccagc 22800 ccatgagccg tcaggtggtg gatgatacta aatacaagga ctaccaacag gtgggcatcc 22860 tacaccaaca caacaactct ggatttgttg gctaccttgc ccccaccatg cgcgaaggac 22920 aggcctaccc tgctaacttc ccctatccgc ttataggcaa gaccgcagtt gacagcatta 22980 cccagaaaaa gtttctttgc gatcgcaccc tttggcgcat cccattctcc agtaacttta 23040 tgtccatggg cgcactcaca gacctgggcc aaaaccttct ctacgccaac tccgcccacg 23100 cgctagacat gacttttgag gtggatccca tggacgagcc cacccttctt tatgttttgt 23160 ttgaagtctt tgacgtggtc cgtgtgcacc agccgcaccg cggcgtcatc gaaaccgtgt 23220 acctgcgcac gcccttctcg gccggcaacg ccacaacata aagaagcaag caacatcaac 23280 aacagctgcc gccatgggct ccagtgagca ggaactgaaa gccattgtca aagatcttgg 23340 ttgtgggcca tattttttgg gcacctatga caagcgcttt ccaggctttg tttctccaca 23400 caagctcgcc tgcgccatag tcaatacggc cggtcgcgag actgggggcg tacactggat 23460 ggcctttgcc tggaacccgc actcaaaaac atgctacctc tttgagccct ttggcttttc 23520 tgaccagcga ctcaagcagg tttaccagtt tgagtacgag tcactcctgc gccgtagcgc 23580 cattgcttct tcccccgacc gctgtataac gctggaaaag tccacccaaa gcgtacaggg 23640 gcccaactcg gccgcctgtg gactattctg ctgcatgttt ctccacgcct ttgccaactg 23700 gccccaaact cccatggatc acaaccccac catgaacctt attaccgggg tacccaactc 23760 catgctcaac agtccccagg tacagcccac cctgcgtcgc aaccaggaac agctctacag 23820 cttcctggag cgccactcgc cctacttccg cagccacagt gcgcagatta ggagcgccac 23880 ttctttttgt cacttgaaaa acatgtaaaa ataatgtact agagacactt tcaataaagg 23940 caaatgcttt tatttgtaca ctctcgggtg attatttacc cccacccttg ccgtctgcgc 24000 cgtttaaaaa tcaaaggggt tctgccgcgc atcgctatgc gccactggca gggacacgtt 24060 gcgatactgg tgtttagtgc tccacttaaa ctcaggcaca accatccgcg gcagctcggt 24120 gaagttttca ctccacaggc tgcgcaccat caccaacgcg tttagcaggt cgggcgccga 24180 tatcttgaag tcgcagttgg ggcctccgcc ctgcgcgcgc gagttgcgat acacagggtt 24240 gcagcactgg aacactatca gcgccgggtg gtgcacgctg gccagcacgc tcttgtcgga 24300 gatcagatcc gcgtccaggt cctccgcgtt gctcagggcg aacggagtca actttggtag 24360 ctgccttccc aaaaagggcg cgtgcccagg ctttgagttg cactcgcacc gtagtggcat 24420 caaaaggtga ccgtgcccgg tctgggcgtt aggatacagc gcctgcataa aagccttgat 24480 ctgcttaaaa gccacctgag cctttgcgcc ttcagagaag aacatgccgc aagacttgcc 24540 ggaaaactga ttggccggac aggccgcgtc gtgcacgcag caccttgcgt cggtgttgga 24600 gatctgcacc acatttcggc cccaccggtt cttcacgatc ttggccttgc tagactgctc 24660 cttcagcgcg cgctgcccgt tttcgctcgt cacatccatt tcaatcacgt gctccttatt 24720 tatcataatg cttccgtgta gacacttaag ctcgccttcg atctcagcgc agcggtgcag 24780 ccacaacgcg cagcccgtgg gctcgtgatg cttgtaggtc acctctgcaa acgactgcag 24840 gtacgcctgc aggaatcgcc ccatcatcgt cacaaaggtc ttgttgctgg tgaaggtcag 24900 ctgcaacccg cggtgctcct cgttcagcca ggtcttgcat acggccgcca gagcttccac 24960 ttggtcaggc agtagtttga agttcgcctt tagtcgtta tccacgtggt acttgtccat 25020. ttggtcaggc cagcgcgcgc gcagcctcca tgcccttctc ccacgcagac acgatcggca cactcagcgg 25080 gttcatcacc gtaatttcac tttccgcttc gctgggctct tcctcttcct cttgcgtccg 25140 cataccacgc gccactgggt cgtcttcatt cagccgccgc actgtgcgct tacctccttt 25200 gccatgcttg attagcaccg gtgggttgct gaaacccacc atttgtagcg ccacatcttc 25260 tctttcttcc tcgctgtcca cgattacctc tggtgatggc gggcgctcgg gcttggggaga 25320 agggcgcttc tttttcttct tgggcgcaat ggccaaatcc gccgccgagg tcgatggccg 25380 cgggctgggt gtgcgcggca ccagcgcgtc ttgtgatgag tcttcctcgt cctcggactc 25440 gatacgccgc ctcatccgct tttttggggg cgcccgggga ggcggcggcg acggggacgg 25500 ggacgacacg tcctccatgg ttgggggacg tcgcgccgca ccgcgtccgc gctcgggggt 25560 ggtttcgcgc tgctcctctt cccgactggc catttccttc tcctataggc agaaaaagat 25620 25680. catggagtca gtcgagaaga aggacagcct aaccgccccc tctgagttcg ccaccaccgc ctccaccgat gccgccaacg cgcctaccac cttccccgtc gaggcacccc cgcttgagga 25740 ggaggaagtg attatcgagc aggacccagg ttttgtaagc gaagacgacg aggaccgctc 25800 agtaccaaca gaggataaaa agcaagacca ggacaacgca gaggcaaacg aggaacaagt 25860 cgggcggggg gacgaaaggc atggcgacta cctagatgtg ggagacgacg tgctgttgaa 25920 gcatctgcag cgccagtgcg ccattatctg cgacgcgttg caagagcgca gcgatgtgcc 25980 cctcgccata gcggatgtca gccttgccta cgaacgccac ctattctcac cgcgcgtacc 26040 ccccaaacgc caagaaaacg gcacatgcga gcccaacccg cgcctcaact tctaccccgt 26100 atttgccgtg ccagaggtgc ttgccaccta tcacatcttt ttccaaaact gcaagatacc 26160 cctatcctgc cgtgccaacc gcagccgagc ggacaagcag ctggccttgc ggcagggcgc 26220 tgtcatacct gatatcgcct cgctcaacga agtgccaaaa atctttgagg gtcttggacg 26280 cgacgagaag cgcgcggcaa acgctctgca acaggaaaac agcgaaaatg aaagtcactc 26340 tggagtgttg gtggaactcg agggtgacaa cgcgcgccta gccgtactaa aacgcagcat 26400 cgaggtcacc cactttgcct acccggcact taacctaccc cccaaggtca tgagcacagt 26460 catgagtgag ctgatcgtgc gccgtgcgca gcccctggag agggatgcaa atttgcaaga 26520 acaaacagag gagggcctac ccgcagttgg cgacgagcag ctagcgcgct ggcttcaaac 26580 gcgcgagcct gccgacttgg aggagcgacg caaactaatg atggccgcag tgctcgttac 26640 cgtggagctt gagtgcatgc agcggttctt tgctgacccg gagatgcagc gcaagctaga 26700 ggaaacattg cactacacct ttcgacaggg ctacgtacgc caggcctgca agatctccaa 26760 cgtggagctc tgcaacctgg tctcctacct tggaattttg cacgaaaacc gccttgggca 26820 aaacgtgctt cattccacgc tcaagggcga ggcgcgccgc gactacgtcc gcgactgcgt 26880 ttacttattt ctatgctaca cctggcagac ggccatgggc gtttggcagc agtgcttgga 26940 ggagtgcaac ctcaaggagc tgcagaaact gctaaagcaa aacttgaagg acctatggac 27000 ggccttcaac gagcgctccg tggccgcgca cctggcggac atcattttcc ccgaacgcct 27060 gcttaaaacc ctgcaacagg gtctgccaga cttcaccagt caaagcatgt tgcagaactt 27120 taggaacttt atcctagagc gctcaggaat cttgcccgcc acctgctgtg cacttcctag 27180 cgactttgtg cccattaagt accgcgaatg ccctccgccg ctttggggcc actgctacct 27240 tctgcagcta gccaactacc ttgcctacca ctctgacata atggagaacg tgagcggtga 27300 cggtctactg gagtgtcact gtcgctgcaa cctatgcacc ccgcaccgct ccctggtttg 27360 caattcgcag ctgcttaacg aaagtcaaat tatcggtacc tttgagctgc agggtccctc 27420 gcctgacgaa aagtccgcgg ctccgggggtt gaaactcact ccggggctgt ggacgtcggc 27480 ttaccttgc aaatttgtac ctgaggacta ccacgcccac gagattaggt tctacgaaga 27540 ccaatcccgc ccgcctaatg cggagcttac cgcctgcgtc attacccagg gccacattct 27600 tggccaattg caagccatca acaaagcccg ccaagagttt ctgctacgaa agggacgggg 27660 ggtttacttg gacccccagt ccggcgagga gctcaaccca atccccccgc cgccgcagcc 27720 ctatcagcag cagccgcggg cccttgctc ccaggatggc acccaaaag aagctgcagc 27780 tgccgccgcc acccacggac gaggagat actgggacag tcaggcag gaggttttgg 27840 27900 27900 27900 tcgaagaggt gtcagacgaa acccgtcac cctcggtcgc attcccctcg ccggcgcccc 27960 agaaatcggc aaccggttcc agcatggcta CAcctccgc tcctcaggcg ccgccggcac 28020 tgcccgttcg ccgacccaac cgtagatggg acaccactgg aaccaggggcc ggtaagtcca 28080 agcagccgcc gccgttagcc caagcaac aacagcgcca aggctaccgc tcatggcgcg 28140 ggcacaagaa cgccatagtt gcttgcttgc aagactgtgg gggcaacatc tccttcgccc 28200 gccgctttct tctctaccat cacggcgtgg ccttcccccg taacacctg cattactacc 28260 gtcatctcta cagcccatac tgcaccggcg gcagcggcag cacagcagc ggccacacag 28320 aaagcaaggc gaccgatag caagactctg aaagcccca agaatccac agcggcggca 28380 gcagcaggag gaggagcgct gcgtctggcg cccaacgaac ccgtatcgac ccgcgagctt 28440 agaaacagga tttttcccac tctgtatgct atatttcaac agagcagggg ccaagaacaa 28500 gagctgaaaa taaaaaacag gtctctgcga tccctcaccc gcagctgcct gtatcacaaa 28560 agcgaagatc agcttcggcg cacgctggaa gacgcggagg ctctcttcag taaatactgc 28620 gcgctgactc ttaaggacta gtttcgcgcc ctttctcaaa tttaagcgcg aaaactacgt 28680 catctccagc ggccacaccc ggcgccagca cctgttgtca gcgccattat gagcaaggaa 28740 attcccacgc cctacatgtg gagttaccag ccacaaatgg gacttgcggc tggagctgcc 28800 caagactact caacccgaat aaactacatg agcgcgggac cccacatgat atcccgggtc 28860 aacggaatac gcgcccaccg aaaccgaatt ctcctggaac aggcggctat taccaccaca 28920 cctcgtaata accttaatcc ccgtagttgg cccgctgccc tggtgtacca ggaaagtccc 28980 gctcccacca ctgtggtact tcccagagac gcccaggccg aagttcagat gactaactca 29040 ggggcgcagc ttgcgggcgg ctttcgtcac agggtgcggt cgcccgggca gggtataact 29100 cacctgacaa tcagagggcg aggtattcag ctcaacgacg agtcggtgag ctcctcgctt 29160 ggtctccgtc cggacgggac atttcagatc ggcggcgccg gccgctcttc attcacgcct 29220 cgtcaggcaa tcctaactct gcagacctcg tcctctgagc cgcgctctgg aggcattgga 29280 actctgcaat ttattgagga gtttgtgcca tcggtctact ttaacccctt ctcgggacct 29340 cccggccact atccggatca atttattcct aactttgacg cggtaaagga ctcggcggac 29400 ggctacgact gaatgttaag tggagaggca gagcaactgc gcctgaaaca cctggtccac 29460 tgtcgccgcc acaagtgctt tgcccgcgac tccggtgagt tttgctactt tgaattgccc 29520 gaggatcata tcgagggccc ggcgcacggc gtccggctta ccgcccaggg agagcttgcc 29580 cgtagcctga ttcgggagtt tacccagcgc cccctgctag ttgagcggga caggggaccc 29640 tgtgttctca ctgtgatttg caactgtcct aaccctggat tacatcaaga tcttattccc 29700 tttaactaat aaaaaaaaat aataaagcat cacttactta aaatcagtta gcaaatttct 29760 gtccagttta ttcagcagca cctccttgcc ctcctcccag ctctggtatt gcagcttcct 29820 cctggctgca aactttctcc acaatctaaa tggaatgtca gtttcctcct gttcctgtcc 29880 atccgcaccc actatcttca tgttgttgca gatgaagcgc gcaagaccgt ctgaagatac 29940 cttcaacccc gtgtatccat atgacacgga aaccggtcct ccaactgtgc cttttcttac 30000 tcctcccttt gtatccccca atgggtttca agagagtccc cctggggtac tctctttgcg 30060 cctatccgaa cctctagtta cctccaatgg catgcttgcg ctcaaaatgg gcaacggcct 30120 ctctctggac gaggccggca accttacctc ccaaaatgta accactgtga gcccacctct 30180 caaaaaaacc aagtcaaaca taaacctgga aatatctgca cccctcacag ttacctcaga 30240 agccctaact gtggctgccg ccgcacctct aatggtcgcg ggcaacacac tcaccatgca 30300 atcacaggcc ccgctaaccg tgcacgactc caaacttagc attgccaccc aaggacccct 30360 cacagtgtca gaaggaaagc tagccctgca aacatcaggc cccctcacca ccaccgatag 30420 cagtaccctt actatcactg cctcaccccc tctaactact gccactggta gcttgggcat 30480 tgacttgaaa gagcccatttt atacacaaa tggaaaacta ggactaaagt acggggctcc 30540 tttgcatgta acagacgacc taaacacttt gaccgtagca actggtccag gtgtgactat 30600 taataatact tccttgcaaa ctaaagttac tggagccttg ggttttgatt cacaaggcaa 30660 tatgcaactt aatgtagcag gaggactag tattgattct caaacagac gccttatact 30720 tgatgttagt tatccgtttg atgctcaaa ccactaaat ctaagactagzgagggccc 30780 tcttttata aactcagccc acaacttgga tattactac aacaaggcc tttactgtt 30840 tacagcttca aacaattcca aaagcttga ggttaaccta agcactgcca aggggttgat 30900 gtttgacgct acagccatag ccattaatgc aggagatggg cttgaatttg gttcacctaa 30960 tgcaccaaac acaaatcccc tcaaaacaa aattggccat ggcctagaat ttgattcaa 31020 caggctatg gttcctaac taggactgg ccttagtttt gatagcacag gtgccattac 31080 agtaggaaac aaaataatg ataagctaac ttgtggacc acaccagctc catctcctaa 31140 ctgtagacta atgcagaga aagatgctaa actcactttg gtctttacaa atgtggcag 31200 tcaatactt gctacagttt cagttttggc tgttaaggc agttttggctc CAATCTgg 31260 aacagttcaa agtgctcatc ttatttaag atttgacgaa atggagtgc tactaacaa 31320 ttccttcctg gacccagaat attggaactt tagaatgga gatcttactg aaggcacagc 31380 ctatacaaac gctgttggat ttgcctaa cctatcagct tatccaaat ctcacggtaa 31440 aactgccaaa agtaacattg tcagtcaagt ttacttaac ggagacaaaa ctaaacctgt 31500 aacactaacc attackacta acggtacaca ggaaacagga vakacaacc caagtgcata 31560 ctctatgtca tttcatggg actggtctgg ccacaacac attaatgaaa tatttgccac 31620 atcctcttac acttttcat acattgccca agaataaga atcgtttgtg ttatgttttca 31680 acgtgtttat ttcattg cagaaaattt caagtcatttt ttcattcagt agtatagccc 31740 caccaccaca tagcttatac agatcaccgt accttaatca aactcacaga accctagtat 31800 tcaacctgcc acctccctcc caccacag agtacacagt ccttctccc cggctggcct 31860 taaaaagcat catatcatgg gtaacagaca tattcttagg tgttatattc cacacggttt 31920 cctgtcgagc caacgctca tcagtgatat taataactc cccgggcagc tcacttaagt 31980 tcatgtcgct gtccagctgc tgagccacag gctgctgtcc aacttgcggt tgcttaacgg 32040 gcggcgaagg agaagtccac gcctacatgg gggtagagtc ataatcgtgc atcaggatag 32100 ggcggtggtg ctgcagcagc gcgcgaataa actgctgccg ccgccgctcc gtcctgcagg 32160 atacaacat ggcagtggtc tcctcagcga tgattcgcac cgcccgcagc ataggcgcc 32220 ttgtcctccg ggcacagcag cgcaccctga tctcacttaa atcagcacag taactgcagc 32280 acagcaccac atattgttc aaatcccac agtgcaaggc gctgtatcca aagctcatgg 32340 cggggaccac agaacccacg tggccatcat accacaagcg caggtagatt aagtggcgac 32400 ccctcataaa cacgctggac aaacatta cctctttgg catgttgtaa ttcaccacct 32460 cccggtacca tataaacctc tgattaaca tggcgccatc caccaccatc ctaaaccagc 32520 tggccaaaac ctgcccgccg gctatacact gcagggaacc gggactggaa caatgacagt 32580 ggagagccca ggactcgtaa ccatggatca tcatgctcgt catgatatca atgttggcac 32640 aacacaggca cacgtgcata cacttctcca ggattacaag ctcctcccgc gttagaacca 32700 tatcccagg aacaacccat tcctgaatca gcgtaaatcc caacactgcag ggaagacctc 32760 gcacgtaact cacgttgtgc attgtcaaag tgttacattc gggcagcagc ggatgatcct 32820 ccagtatggt agcgcgggtt tctgtctcaa aaagggtag acgatcccta ctgtacggag 32880 tgcgccgaga caaccgagat cgtgttggtc gtagtgtcat gccaaatgga acgccggacg 32940 tagtcatatt tcctgaagca aaaccaggtg cgggcgtgac aaacagatct gcgtctccgg 33000 tctcgccgct tagatcgctc tgtgtagtag ttgtagtata tccactctct caaagcatcc 33060 aggcgccccc tggcttcggg ttctatgtaa actccttcat gcgccgctgc cctgataaca 33120 tccaccaccg cagaataagc caacccagc caacctacac attcgttctg cgagtcacac 33180 acgggagg cgggaagagc tggagaaacc atgttttttt ttttattcca aaagattatc 33240 caaaacctca aaatgaagat ctattaagtg aacgcgctcc cctccggtgg cgtggtcaaa 33300 ctctacagcc aaagaacaga taatggcatt tgtaagatgt tgcacaatgg cttccaaaag 33360 gcaaacggcc ctcacgtcca agtggacgta aaggctaaac ccttcagggt gaatctcctc 33420 tataaacatt ccagcacctt caaccatgcc caaataattc tcatctcgcc accttctcaa 33480 tatatctcta agcaaatccc gaatattaag tccggccatt gtaaaaatct gctccagagc 33540 gccctccacc ttcagcctca agcagcgaat catgattgca aaaattcagg ttcctcacag 33600 acctgtataa gattcaaaag cggaacatta acaaaaatac cgcgatcccg taggtccctt 33660 cgcagggcca gctgaacata atcgtgcagg tctgcacgga ccagcgcggc cacttccccg 33720 ccaggaacca tgacaaaaga acccacactg attatgacac gcatactcgg agctatgcta 33780 accagcgtag ccccgatgta agcttgttgc atgggcggcg atataaaatg caaggtgctg 33840 ctcaaaaaat caggcaaagc ctcgcgcaaa aaagaaagca catcgtagtc atgctcatgc 33900 agataaaggc aggtaagctc cggaaccacc acagaaaaag acaccatttt tctctcaaac 33960 atgtctgcgg gttctctgcat aaaacaaaa taaaataca aaaaaacatt taaacattag 34020 aagcctgtct tacacagga aaaacaaccc ttataagcat aagacggact acggccatgc 34080 cggcgtgacc gtaaaaaaac tggtcaccgt gattaaaag caccaccgac agctcctcgg 34140 tcatgtccgg agtcataatg tagactcgg taaacacatc agttgattc acatcggtca 34200 gtgctaaaaa gcgaccgaaa tagcccgggg gatacatac ccgcaggcgt agagacaaca 34260 ttacagcccc cataggaggt atacaaat tataggaga gaaaacaca taacaccctg 34320 aaaaaccctc cctccctaggc aaatagcac cctcccgctc cagacaaca tacagcgctt 34380 ccacagcggc agccataca gtcagcctta ccagtaaaaa agaaaaccta ttaaaaaaac 34440 accactcgac acggcaccag ctcatcagt cacagtgtaa aaagggcca agtgcagagc 34500 gagtatatat aggactaaa atgacgtaa cggttaaagt ccacaaaaaa caccagaaa 34560 accgcacgcg aacctacgcc cagaaacgaa agccaaaaa cccacaactt cctcaatcg 34620 tcacttccgt tttcccacgt tacgtcactt cccattttaa gaaaactaca attcccaaca 34680 catacaagtt actccgccct aaacctacg tcacccgccc cgttcccacg cccgcgcca 34740 cgtcacaaac tccaccccct cattatcata ttggcttcaa tccaaaataa ggtatattat 34800 tgatgatg 34808 <210> 7 <211> 34742 <212> DNA <213> Recombinant human adenovirus serotype 5 <400> 7 ccatcttcaa tatatacct caaacttttt tgtgcgcgtt aatatgcaaa tgaggcgttt 60 gaatttgggg aggaagggcg gtgattggtc gagggatgag cgaccgttag gggcggggcg 120 agtgacgttt tgatgacgtg gttgcgagga ggagccagtt tgcaagttct cgtgggaaaa 180 gtgacgtcaa acgaggtgtg gtttgaacac ggaaatactc aatttcccg cgctctctga 240 caggaaatga ggtgtttctg ggcggatgca agtgaaaacg ggccattttc gcgcgaaaac 300 tgaatgagga agtgaaaatc tgagtaattt cgcgtttatg gcagggagga gtatttgccg 360 agggccgagt agactttgac cgattacgtg ggggtttcga ttaccgtgtt tttcacctaa 420 atttccgcgt acggtgtcaa agtccggtgt tttacgtag gtgtcagctg atcgccaggg 480 tatttaaacc tgcgctctcc agtcaagagg ccactcttga gtgccagcga gaagagttgc 540 gatgcatgcg cgtgtcaaa tatgagctca caatgcttc atcaaacgag ttggtgctca 600 tggcggcggc ggctgctgca aaacagatac aaaactacat aagaccccca ccttatatat 660 tctttcccac ccttaaccac gcccagatcc gcgttaagat acattgatga gtttggacaa 720 accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct 780 ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg cattcatttt 840 atgtttcagg ttcagggggga ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaaa 900 tgtggtatgg ctgattatga tcagttatct agactcgagc ggccgcgata tcttatgtgt 960 aatgtaattt gactcctttg agcacgggct cagaatcgtc ctcgtcgaac ttgcagcagc 1020 tgccacagct acaacagcccc ttcaggcagc tacagcagct ggtcatgcaa cacagcatga 1080 ttgtgaccat cacgatggca atcagtccgg cgataaagcc cagccagatg taccagggcc 1140 acttgatgta ctgctcgtac ttccccagtt cttgcaggtc gatcaggctc tcgttcagat 1200 tcttggccac ctcgttcagc cggtcgatct ctttctggat gttcacgacg ctggcattga 1260 ttccgctgat gtcgcccagg tccacgtcgg ggcttgtgtg gttcttaaag tacttatcca 1320 gttcctcttt gaagctgtcc agctcgggct gcagagggtc gtacacggta ttgttcacaa 1380 tgccgatcac gacgtcgcag ttgccagaca cgaaggtgtt gtcggtggtg atgatctggg 1440 gctcgtagaa gttccgctgg gtcacgaacc aatgggtgcc gttggacacg aacacgcctt 1500 ctctaggaaa gtgggctttg ccgtcgtggc agatggctgg agcggtggtg aaattcttct 1560 cttgagcggg cacgtatgtc acgtgcagaa acaccacgcc gtgaggggca gactgaggga 1620 agctcatcag gtggtagccc ttgccgcaaa agtccactct cttgctctgg cccagcacac 1680 actcagacat cttggtggcg gccagattgg cagaggctct aatctcggcg gctctgatca 1740 gctgctgggt aacgtaggtc tgcagggact gcagccttcc ggtgatcagt ctgtcgatct 1800 gcacctcggc ttccaccttg tccagtctgc tcaggatgtc gttcagcaca gagctgatgg 1860 cgccgaagtt ggaggacagc tgcttgacca gggtgttcag tgcctgggca ttctggttga 1920 ccacgtcctg cagctttccc agggcgcttg ctgtgctgct caggctgtcc tggatcttgc 1980 cgatggcgct gttgaactgg ttggcgatca gcttctggtt ctcgtacagc acattctggg 2040 tcactccgat gccgttgaac cggtaggcca tctgcatagc aaaggggatc tgcagagcgg 2100 cgccagctcc aaatgtccag ccgcttgtga ttgtgccggc cagcagggca gatgtgtact 2160 gggcgatcat ctcatcggtc agcagtggtg gcagcactgt cagtccgtta aacttctggg 2220 cgcaaatcag atccctggcg gcaatgtcgc ccagacaatc gccatactgc ttgatgaagc 2280 cggcgtcggc cagtgtcact ttgttgaaca gcaggtcctc gatgaagctc cgcttgctgg 2340 gcttgctagg atcgggcaga atctggctga aattgaagcc gccgaagtcc ttgataggag 2400 gggtcttgta gatctgcttc acttgggcga acacctcttg ggtgttcttg tcctgttcca 2460 cggcgatccc tgtcagggct ctattcagct gggtgcagaa gctgccgtac tgcagcagca 2520 ggttggagca ctcggtggaa tcgccgcaga tgtacatggt gcagtccacg ctggtcttgg 2580 tcatggacac aggcaggatc tctgtggtca cgctgatggt gaagttggtg gggatagcga 2640 tagagttgtt ggagtaggcc acgctgttct cggcgcccag agacattgtg taggcaatga 2700 tgctctggct ggccacagat ctggcccgtc tggggctgtt tgtctgtgtc tggtagctgg 2760 cacagatgcc agcgccgatg gggatgtcgc actcgtagct attgttcacg tgctcggctc 2820 cgatcagaca gccggctctg gtctgaaaca cattgctgcc ggtggagtac acccgccatg 2880 taggtgtcag ctgatcggcg tgaatggcca cgggcacttc ggtacagttc acgtcctggt 2940 acagcactgc cacctgattg ctggtgttgg tgccaggggt gatcacagac actccgccga 3000 agctgcaagg ggtgatgtcc aggatttcca gtgtctgggg atctcgtacg gcgtctgtgg 3060 tatcggcaat atcccggcca aactgctgga atggcaggaa cttcttgttg ctctctgtca 3120 gcacgccggt gccggtcagg ccgttgaagt tgaagttcac gcatttgttc ttcacgagat 3180 tggtgctttt cttagggccg cacactgtgg caggggcatg cagcagttcg aagctcagca 3240 ccaccactct gtagggctga tagcccacgc catttgtggg ctgaaagccg taggactgca 3300 gtgggagta gcagttgaag ccttccacgc cgttacaagg ggtgctgccg gcctgataga 3360 tctcggtgga gatgtcccgc tcgaagggct tcagattgga cttccggac agccggtaca ggtaattgta gttgccgccg actttggagt ccaggttgtt gctgttccag gcaatcacac 3480 agccggtgaa gtcgtcgggc agcttgtagt tgtagtcggc gatcttgcct gtctgtccag 3540 gggcaatctg ccgcacttca tctccccgga tcacgaagct gtcggcgtac acgtttgtga 3600. agcacaggtc gttcagcttg gtaggggaca cgccgtagca cttgaaggtg ctgaagctgg cggagttgta cagcacggag tagtcggcca cgcaattgct gatccgcttc cggttccagg 3720 cgtacacaga ggcgaatctg gtggcattga acacctcgcc gaaggggcac agattggtga tattgggga ccgcacgatg gattcggtgg gctgcacccg gagttgctg gtctggtaga 3840 tgcccttttc cacggtgag gacttcaggg tgcactttgt ctcgctcaga ggatccagag cacaatccac ggcgtcggtg atggtgccgt tctcgttgta cttcagcagg aaggttctag gctgcaggta gcccacatag tagcggcgg caccagctgt ccatccgctg ctgctatcgc 4020 caggtgtcag gtagcttctg tgcagggcca gcagtgtctg aaaccgggtg atgttgatgc 4080 cgatgggcag atccaccagg ggttccagag cagagaagcc ctgaggcaga tccccgcacga ggttgatagg ggtgtgcttg ctgtagatct tgaagtagcc gtcgatgttc ttgaacacga actcgcgcag gttcttgag ttgccctgct tgccttccag gtccatcagg aaaggctggg 4260 acacgtactc gaaggtgcag ttgttggcgc tgctgtacac ccggaactcg ctttccatcc agctcttgtt gttcttgtga tagtaggc ccaggaaggg gtcgttgcag aactggaact 4380 cgcacacttt gatgaccacg ttggtggcgt tgttcacgat cagcaggctc tgggtcttgc 4440 tgtccagtgt ggtgccgaag atccagcctc tgatgatgtt ggacttctcg gtgctggcaa agtacacccc gtcgttgaag ggcagcacgg ggttgtcgaa tctcttggtg ccattggtgc 4560. cggacacgtg gatggcgtgg aaccaggtca cgttgctgaa gaaaggcagg aacaggtcct gggtaggtg cagcacgctg gatctgaaca ccttgtcggg gtagtacacg cctctggtaa agctgttggt gtaggctgga ggcagctggg ttcttgtggt caggttcaca cactgactag 4740 agactagtgg caataacaca aggaacacaa acatggtacc agatctctag cggatctgac 4800 ggttcactaa accagctctg cttatataga cctcccaccg tacacgccta ccgcccattt 4860 gcgtcaatgg ggcggagttg ttacgacatt ttggaaagtc ccgttgattt tggtgccaaa 4920 acaaactccc attgacgtca atggggtgga gacttggaaa tccccgtgag tcaaaccgct 4980 atccacgccc attgatgtac tgccaaaacc gcatcaccat ggtaatagcg atgactaata 5040 cgtagatgta ctgccaagta ggaaagtccc ataaggtcat gtactgggca taatgccagg 5100 cgggccattt accgtcattg acgtcaatag ggggcgtact tggcatatga tacacttgat 5160 gtactgccaa gtgggcagtt taccgtaaat actccaccca ttgacgtcaa tggaaagtcc 5220 ctattggcgt tactatggga acatacgtca ttattgacgt caatgggcgg gggtcgttgg 5280 gcggtcagcc aggcgggcca tttaccgtaa gttatgtaac gcggaactcc atatatgggc 5340 tatgaactaa tgaccccgta attgattact attacagtat tacgcgctat gagtaacaca 5400 aaattattca gatttcactt cctcttattc agttttcccg cgaaaatggc caaatcttac 5460 5520. tcggttacgc ccaaatttac tacaacatcc gcctaaaacc gcgcgaaaat tgtcacttcc tgtgtacacc ggcgcgctga gtagtgttct ggggcgggggg aggacctgca tgagggccag 5580. aataactgaa atctgtgctt ttctgtgtgt tgcagcagca tgagcggaag cggctccttt gagggagggg tattcagccc ttatctgacg gggcgtctcc cctcctgggc gggagtgcgt 5700 cagaatgtga tgggatccac ggtggacggc cggcccgtgc agcccgcga ctcttcaacc ctgacctatg caaccctgag ctcttcgtcg ttggacgcag ctgccgccgc agctgctgca 5820. tctgccgcca gcgccgtgcg cggaatggcc atgggcgccg gctactacgg cactctggtg 5880 gccaactcga gttccacca taatccccgcc agcctgaacg aggagaagct gttgctgctg atggcccagc tcgaggcctt gacccagcgc ctgggcgagc tgacccagca ggtggctcag ctgcaggagc agacgcgggc cgcggttgcc acggtgaaat ccaataaa aatgaatcaa father gagacggttg ttgattttaa cacagagtct gaatctttat ttgatttttc gcgcgcggta ggccctggac caccggtctc gatcattgag cacccggtgg atcttttcca 6180. ggacccggta gaggtgggct tggatgttga ggtacatggg catgagcccg tcccgggggt 6240 ggaggtagct ccattgcagg gcctcgtgct cgggggtggt gttgtaaatc acccagtcat 6300 agcaggggcg cagggcatgg tgttgcacaa tatctttgag gaggagactg atggccacgg 6360 gcagcccttt ggtgtaggtg tttacaaatc tgttgagctg ggagggatgc atgcgggggg 6420 agatgaggtg catcttggcc tggatcttga gattggcgat gttaccgccc agatcccgcc 6480 tggggttcat gttgtgcagg accaccagca cggtgtatcc ggtgcacttg gggaatttat 6540 catgcaactt ggaagggaag gcgtgaaaga atttggcgac gcctttgtgc ccgcccaggt 6600 tttccatgca ctcatccatg atgatggcga tgggcccgtg ggcggcggcc tgggcaaaga 6660 cgtttcgggg gtcggacaca tcatagttgt ggtcctgggt gaggtcatca taggccattt 6720 taatgaattt ggggcggagg gtgccggact gggggacaaa ggtaccctcg atcccggggg 6780 cgtagttccc ctcacagatc tgcatctccc aggctttgag ctcggagggg gggatcatgt 6840 ccacctgcgg ggcgataaag aacacggttt ccggggcggg ggagatgagc tgggccgaaa 6900 gcaagttccg gagcagctgg gacttgccgc agccggtggg gccgtagatg accccgatga ccggctgcag gtggtagttg agggagac agctgccgtc ctcccggagg agggggcca 7020 cctcgttcat catctcgcgc acgtgcatgt tctcgcgcac cagttccgcc aggaggcgct 7080 ctccccccag ggataggagc tcctggagcg aggcgagtt tttcagcggc ttgagtccgt 7140 cggccatggg cattttggag agggtttgtt gcaagagttc caggcggtcc cagagctcgg 7200. tgatgtgctc tacggcatct cgatccagca gacctcctcg tttcgcgggt tgggacggct 7260. gcgggagtag ggcaccagac gatggggcgtc cagcgcagcc agggtccggt ccttccaggg 7320 tcgcagcgtc cgcgtcaggg tggtctccgt cacggtgaag gggtgcgcgc cgggctgggc 7380. gcttgcgagg gtgcgcttca ggctcatccg gctggtcgaa aaccgctccc gatcggcgcc 7440 ctgcgcgtcg gccaggtagc aattgaccat gagttcgtag ttgagcgcct cggccgcgtg 7500. gccttttggcg cggagcttac ctttggaagt ctgcccgcag gcgggacaga ggagggactt 7560 gagggcgtag agcttggggg cgaggagac ggactcgggg gcgtaggcgt ccgcgccgca 7620. gtgggcgcag acggtctcgc actccacgag ccaggtgagg tcgggctggt cggggtcaaa 7680 aaccagtttc ccgccgttct ttttgatgcg tttcttacct ttggtctcca tgagctcgtg 7740 tccccgctgg gtgacaaaga ggctgtccgt gtccccgtag accgacttta tgggccggtc 7800 ctcgagcggt gtgccgcggt cctcctcgta gaggaacccc gcccactccg agacgaaagc 7860 ccgggtccag gccagcacga aggaggccac gtgggacggg tagcggtcgt tgtccaccag 7920 cgggtccacc ttttccaggg tatgcaaaca catgtccccc tcgtccacat ccaggaaggt 7980 gattggcttg taagtgtagg ccacgtgacc gggggtcccg gccggggggg tataaaaggg 8040 tgcgggtccc tgctcgtcct cactgtcttc cggatcgctg tccaggagcg ccagctgttg 8100 gggtaggtat tccctctcga aggcgggcat gacctcggca ctcaggttgt cagtttctag 8160 aaacgaggag gatttgatat tgacggtgcc ggcggagatg cctttcaaga gcccctcgtc 8220 catctggtca gaaaagacga tctttttgtt gtcgagcttg gtggcgaagg agccgtagag 8280 ggcgttggag aggagcttgg cgatggagcg catggtctgg tttttttcct tgtcggcgcg 8340 ctccttggcg gcgatgttga gctgcacgta ctcgcgcgcc acgcacttcc attcggggaa 8400 gacggtggtc agctcgtcgg gcacgattct gacctgccag ccccgattat gcagggtgat 8460 gaggtccaca ctggtggcca cctcgccgcg caggggctca ttagtccagc agaggcgtcc 8520 gcccttgcgc gagcagaagg ggggcagggg gtccagcatg acctcgtcgg gggggtcggc 8580 atcgatggtg aagatgccgg gcaggaggtc ggggtcaaag tagctgatgg aagtggccag 8640 atcgtccagg gcagcttgcc attcgcgcac ggccagcgcg cgctcgtagg gactgagggg 8700 cgtgccccag ggcatgggat gggtaagcgc ggaggcgtac atgccgcaga tgtcgtagac 8760 gtagaggggc tcctcgagga tgccgatgta ggtggggtag cagcgccccc cgcggatgct 8820 ggcgcgcacg tagtcataca gctcgtgcga gggggcgagg agccccgggc ccaggttggt 8880 gcgactgggc ttttcggcgc ggtagacgat ctggcggaaa atggcatgcg agttggagga 8940 gatggtgggc ctttggaaga tgttgaagtg ggcgtggggc agtccgaccg agtcgcggat 9000 gaagtgggcg taggagtctt gcagcttggc gacgagctcg gcggtgacta ggacgtccag 9060 agcgcagtag tcgaggtct cctggatgat gtcatacttg agctgtccct tttgtttcca 9120 cagctcgcgg ttgagaagga actcttcgcg gtccttccag tactcttcga gggggaaccc 9180 gtcctgatct gcacggtaag agcctagcat gtagaactgg ttgacggcct tgtaggcgca 9240 gcagcccttc tccacgggga gggcgtaggc ctgggcggcc ttgcgcagg aggtgtgcgt 9300 gagggcgaaa gtgtccctga ccatgacctt gaggaactgg tgcttgaagt cgatatcgtc 9360 gcagcccccc tgctcccaga gctggaagtc cgtgcgcttc ttgtaggcgg ggttgggcaa 9420 agcgaaagta acatcgttga agaggatctt gcccgcgcgg ggcataaagt tgcgagtgat 9480 gcggaaaggt tggggcacct cggcccggtt gttgatgacc tgggcggcga gcacgatctc 9540 gtcgaagccg ttgatgttgt ggcccacgat gtagagttcc acgaatcgcg gacggccctt 9600 gacgtggggc agtttcttga gctcctcgta ggtgagctcg tcggggtcgc tgagcccgtg 9660 ctgctcgagc gcccagtcgg cgagatgggg gttggcgcgg aggaaggaag tccagagatc 9720 cacggccagg gcggtttgca gacggtcccg gtactgacgg aactgctgcc cgacggccat 9780 tttttcgggg gtgacgcagt agaaggtgcg ggggtccccg tgccagcgat cccatttgag 9840 ctggagggcg agatcgaggg cgagctcgac gagccggtcg tccccggaga gtttcatgac 9900 cagcatgaag gggacgagct gcttgccgaa ggaccccatc caggtgtagg tttccacatc 9960 gtaggtgagg aagagccttt cggtgcgagg atgcgagccg atggggaaga actggatctc 10020 ctgccaccaa ttggaggaat ggctgttgat gtgatggaag tagaaatgcc gacggcgcgc 10080 cgaacactcg tgcttgtgtt tatacaagcg gccacagtgc tcgcaacgct gcacgggatg 10140 cacgtgctgc acgagctgta cctgagttcc tttgacgagg aatttcagtg ggaagtggag 10200 tcgtggcgcc tgcatctcgt gctgtactac gtcgtggtgg tcggcctggc cctcttctgc 10260 ctcgatggtg gtcatgctga cgagcccgcg cgggaggcag gtccagacct cggcgcgagc 10320 gggtcggaga gcgaggacga gggcgcgcag gccggagctg tccagggtcc tgagacgctg 10380 cggagtcagg tcagtgggca gcggcggcgc gcggttgact tgcaggagtt tttccagggc 10440 gcgcgggagg tccagatggt acttgatctc caccgcgcca ttggtggcga cgtcgatggc 10500 ttgcagggtc ccgtgcccct ggggtgtgac caccgtcccc cgtttcttct tgggcggctg 10560 gggcgacggg ggcggtgcct cttccatggt tagaagcgg ggcgaggacg cgcgccggc 10620 ggcaggggcg gctcggggcc cggaggcagg ggcggcaggg gcacgtcggc gccgcgcgcg 10680 ggtaggttct ggtactgcgc ccggagaaga ctggcgtgag cgacgacgcg acggttgacg 10740 tcctggatct gacgcctctg ggtgaaggcc acgggacccg tgagtttgaa cctgaaagag 10800 agttcgacag aatcaatctc ggtatcgttg acggcggcct gccgcaggat ctcttgcacg 10860 tcgcccgagt tgtcctggta ggcgatctcg gtcatgaact gctcgatctc ctcctcttga 10920 aggtctccgc ggccggcgcg ctccacggtg gccgcgaggt cgttggagat gcggcccatg 10980 agctgcgaga aggcgttcat gcccgcctcg ttccagacgc ggctgtagac cacgacgccc 11040 tcgggatcgc cggcgcgcat gaccacctgg gcgaggttga gctccacgtg gcgcgtgaag 11100 accgcgtagt tgcagaggcg ctggtagagg tagttgagcg tggtggcgat gtgctcggtg 11160 acgaagaaat acatgatcca gcggcggagc ggcatctcgc tgacgtcgcc cagcgcctcc 11220 aaacgttcca tggcctcgta aaagtccacg gcgaagttga aaaactggga gttgcgcgcc 11280 gagacggtca actcctcctc cagaagacgg atgagctcgg cgatggtggc gcgcacctcg 11340 cgctcgaagg cccccgggag ttcctccact tcctcttctt cctcctccac taacatctct 11400 tctacttcct cctcaggcgg cagtggtggc gggggagggg gcctgcgtcg ccggcggcgc 11460 acgggcagac ggtcgatgaa gcgctcgatg gtctcgccgc gccggcgtcg catggtctcg 11520 gtgacggcgc gcccgtcctc gcggggccgc agcgtgaaga cgccgccgcg catctccagg 11580 tggccggggg ggtccccgtt gggcagggag agggcgctga cgatgcatct tatcaattgc 11640 cccgtaggga ctccgcgcaa ggacctgagc gtctcgagat ccacgggatc tgaaaaccgc 11700 tgaacgaagg cttcgagcca gtcgcagtcg caaggtaggc tgagcacggt ttcttctggc 11760 gggtcatgtt ggttgggagc ggggcgggcg atgctgctgg tgatgaagtt gaaataggcg 11820 gttctgagac ggcggatggt ggcgaggagc accaggtctt tgggcccggc ttgctggatg 11880 cgcagacggt cggccatgcc ccaggcgtgg tcctgacacc tggccaggtc cttgtagtag 11940 tcctgcatga gccgctccac gggcacctcc tcctcgcccg cgcggccgtg catgcgcgtg 12000 agcccgaagc cgcgctgggg ctggacgagc gccaggtcgg cgacgacgcg ctcggcgagg 12060 atggcttgct ggatctgggt gagggtggtc tggaagtcat caaagtcgac gaagcggtgg 12120 taggctccgg tgttgatggt gtaggagcag ttggccatga cggaccagtt gacggtctgg 12180 tggcccggac gcacgagctc gtggtacttg aggcgcgagt aggcgcgcgt gtcgaagatg 12240 tagtcgttgc aggtgcgcac caggtactgg tagccgatga ggaagtgcgg cggcggctgg 12300 cggtagagcg gccatcgctc ggtggcgggg gcgccgggcg cgaggtcctc gagcatggtg 12360 cggtggtagc cgtagatgta cctggacatc caggtgatgc cggcggcggt ggtggaggcg 12420 cgcgggaact cgcggacgcg gttccagatg ttgcgcagcg gcaggaagta gttcatggtg 12480 ggcacggtct ggcccgtgag gcgcgcgcag tcgtggatgc tctatacggg caaaaacgaa 12540 agcggtcagc ggctcgactc cgtggcctgg aggctaagcg aacgggttgg gctgcgcgtg 12600 taccccggtt cgaatctcga atcaggctgg agccgcagct aacgtggtat tggcactccc 12660 gtctcgaccc aagcctgcac caaccctcca gcatacggag gcgggtcgtt ttgcaacttt 12720 tttttggagg ccggatgaga ctagtaagcg cggaaagcgg ccgaccgcga tggctcgctg 12780 ccgtagtctg gagaaatc gccagggttg cgttgcggtg tgccccggtt cgaggccggc 12840 cggattccgc ggctaacgag ggcgtggctg ccccgtcgtt tccaagaccc catagccagc 12900 cgacttctcc agttacggag cgagcccctc ttttgttttg tttgtttttg ccagatgcat 12960 cccgtactgc ggcagatgcg ccccaccac cctccaccgc aacaacagcc ccctccacag 13020 ccggcgcttc tgccccccgcc ccagcagcaa cttccagcca cgaccgccgc ggccgccgtg 13080 agcggggctg gacagagtta tgatcaccag ctggccttgg aagagggcga ggggctggcg 13140 cgcctgggg cgtcgtcgcc ggagcggcac ccgcgcgtgc agatgaaaag ggacgctcgc 13200 gaggcctacg tgcccaagca gaacctgttc agagagga gcggcgagga gcccgaggag 13260 atgcgcgcgg cccggttcca cgcggggcgg gagctgcggc gcggcctgga ccgaaagagg 13320 gtgctgaggg acgaggattt cgaggcggac gagctgacgg ggatcagccc cgcgcgcgcg 13380 cacgtggccg cggccaacct ggtcacggcg tacgagcaga ccgtgaagga ggagagcaac 13440 ttccaaaaat ccttcaacaa cccgtgcgc accctgatcg cgcgcgagga ggtgaccctg ggcctgatgc acctgtggga cctgctggag gccatcgtgc agaaccccac cagcaagccg 13560 ctgacggcgc agctgttcct ggtggtgcag catagtcggg acaacgaagc gttcagggag gcgctgctga atatcaccga gcccgagggc cgctggctcc tggacctggt gaacattctg cagagcatcg tggtgcagga gcgcgggctg ccgctgtccg agaagctggc ggccatcaac 13740 ttctcggtgc tgagtttggg caagtactac gctaggaaga tctacaagac cccgtacgtg cccatagaca aggaggtga gatcgacggg ttttacatgc gcatgaccct gaaagtgctg accctgagcg acgatctggg ggtgtaccgc aacgacagga tgcaccgtgc ggtgagcgcc 13920 agcaggcggc gcgagctgag cgaccaggag ctgatgcata gtctgcagcg ggccctgacc 13980 ggggccggga ccgaggggga gagctacttt gacatgggcg cggacctgca ctggcagccc 14040 agccgccggg ccttggaggc ggcggcagga ccctacgtag aagaggtgga cgatgaggtg 14100 gacgagggg gcgagtacct ggaagactga tggcgcgacc gtatttttgc tagatgcaac 14160 aacaacagcc acctcctgat cccgcgatgc gggcggcgct gcagagccag ccgtccggca 14220 ttaactctc ggacgattgg acccaggcca tgcaacgcat catggcgctg acgacccgca 14280 accccgaagc ctttagacag cagcccccagg ccaaccggct ctcggccatc ctggaggccg 14340 tggtgccctc gcgctccaac cccacgcacg agaaggtcct ggccatcgtg aacgcgctgg 14400 tggagaacaa ggccatccgc ggcgacgagg ccggcctggt gtacaacgcg ctgctggagc 14460 gcgtggcccg ctacaacagc accaacgtgc agaccaacct ggaccgcatg gtgaccgacg 14520 tgcgcgaggc cgtggcccag cgcgagcggt tccaccgga gtccaacctg ggatccatgg 14580 tggcgctgaa cgcttctctc agcacccagc ccgccaacgt gccccggggc caggaggact 14640 acaccaactt catcagcgcc ctgcgcctga tggtgaccga ggtgccccag agcgaggtgt 14700 accagtccgg gccggactac ttcttccaga ccagtcgcca gggcttgcag accgtgaacc 14760 tgagccaggc tttcaagaac ttgcagggcc tgtggggcgt gcaggccccg gtcggggacc 14820 gcgcgacggt gtcgagcctg ctgacgccga actcgcgcct gctgctgctg ctggtggccc 14880 ccttcacgga cagcggcagc atcaaccgca actcgtacct gggctacctg attaacctgt 14940 accgcgaggc catcggccag gcgcacgtgg acgagcagac ctaccaggag atcacccacg 15000 tgagccgcgc cctgggccag gacgacccgg gcaacctgga agccaccctg aactttttgc 15060 tgaccaaccg gtcgcagaag atcccgcccc agtacgcgct cagcaccgag gaggagcgca 15120 tcctgcgtta cgtgcagcag agcgtgggcc tgttcctgat gcaggagggg gccaccccca 15180 gcgccgcgct cgacatgacc gcgcgcaaca tggagcccag catgtacgcc agcaaccgcc 15240 cgttcatcaa taaactgatg gactacttgc atcgggcggc cgccatgaac tctgactatt 15300 tcaccaacgc catcctgaat ccccactggc tcccgccgcc ggggttctac acgggcgagt 15360 acgacatgcc cgaccccaat gacgggttcc tgtgggacga tgtggacagc agcgtgttct 15420 ccccccgacc gggtgctaac gagcgcccct tgtggaagaa ggaaggcagc gaccgacgcc 15480 cgtcctcggc gctgtccggc cgcgagggtg ctgccgcggc ggtgcccgag gccgccagtc 15540 ctttcccgag cttgcccttc tcgctgaaca gtatccgcag cagcgagctg ggcaggatca 15600 cgcgcccgcg cttgctgggc gaagagggt acttgaatga ctcgctgttg agacccgagc 15660 gggagaagaa cttccccaat aacgggatag aaagcctggt ggacaagatg agccgctgga 15720 agacgtatgc gcaggagcac agggacgatc cccgggcgtc gcaggggcc acgagccggg 15780 gcagcgccgc ccgtaaacgc cggtggcacg acaggcagcg gggacagatg tgggacgatg 15840 aggactccgc cgacgacagc agcgtgttgg acttgggtgg gagtggtaac ccgttcgctc 15900 acctgcgccc ccgtatcggg cgcatgatgt aagagaaacc gaaaataat gatactcacc 15960 aaggccatgg cgaccagcgt gcgttcgttt cttctctgtt gttgttgtat ctagtatgat 16020 gaggcgtgcg tacccggagg gtcctcctcc ctcgtacgag agcgtgatgc agcaggcgat 16080 ggcggcggcg gcgatgcagc ccccgctgga ggctccttac gtgcccccgc ggtacctggc 16140 gcctacggag gggcggaaca gcattcgtta ctcggagctg gcacccttgt acgataccac 16200 ccggttgtac ctggtggaca acaagtcggc ggacatcgcc tcgctgaact accagaacga 16260 ccacagcaac ttcctgacca ccgtggtgca gaacaatgac ttcaccccca cggaggccag 16320 cacccagacc atcaactttg acgagcgctc gcggtggggc ggccagctga aaaccatcat 16380 gcacaccaac atgcccaacg tgaacgagtt catgtacagc aacaagttca aggcgcgggt 16440 gatggtctcc cgcaagaccc ccaatggggt gacagtgaca gaggattatg atggtagtca 16500 ggatgagctg aagtatgaat gggtggaatt tgagctgccc gaaggcaact tctcggtgac 16560 catgaccatc gacctgatga acaacgccat catcgacaat tacttggcgg tggggcggca 16620 gaacggggtg ctggagagcg acatcggcgt gaagttcgac actaggaact tcaggctggg 16680 ctgggacccc gtgaccgagc tggtcatgcc cggggtgtac accaacgagg ctttccatcc 16740 cgatattgtc ttgctgcccg gctgcggggt ggacttcacc gagagccgcc tcagcaacct 16800 gctgggcatt cgcaagaggc agcccttcca ggaaggcttc cagatcatgt acgaggatct 16860 ggaggggggc aacatccccg cgctcctgga tgtcgacgcc tatgagaaaa gcaaggagga 16920 tgcagcagct gaagcaactg cagccgtagc taccgcctct accgaggtca ggggcgataa 16980 ttttgcaagc gccgcagcag tggcagcggc cgaggcggct gaaaccgaaa gtaagatagt 17040 cattcagccg gtggagaagg atagcaagaa caggagctac aacgtactac cggacaagat 17100 aaacaccgcc taccgcagct ggtacctagc ctacaactat ggcgaccccg agaagggcgt 17160 gcgctcctgg acgctgctca ccacctcgga cgtcacctgc ggcgtggagc aagtctactg 17220 gtcgctgccc gacatgatgc aagacccggt caccttccgc tccacgcgtc aagttagcaa 17280 ctacccggtg gtgggcgccg agctcctgcc cgtctactcc aagagcttct tcaacgagca 17340 ggccgtctac tcgcagcagc tgcgcgcctt cacctcgctt acgcacgtct tcaaccgctt 17400 ccccgagaac cagatcctcg tccgcccgcc cgcgcccacc attaccaccg tcagtgaaaa 17460 cgttcctgct ctcacagatc acgggaccct gccgctgcgc agcagtatcc ggggagtcca 17520 gcgcgtgacc gttactgacg ccagacgccg cacctgcccc tacgtctaca aggccctggg 17580 catagtcgcg ccgcgcgtcc tctcgagccg caccttctaa atgtccattc tcatctcgcc 17640 cagtaataac accggttggg gcctgcgcgc gcccagcaag atgtacggag gcgctcgcca 17700 acgctccacg caacaccccg tgcgcgtgcg cgggcacttc cgcgctccct ggggcgccct 17760 caagggccgc gtgcggtcgc gcaccaccgt cgacgacgtg atcgaccagg tggtggccga 17820 cgcgcgcaac tacacccccg ccgccgcgcc cgtctccacc gtggacgccg tcatcgacag 17880 cgtggtggcg gacgcgcgcc ggtacgcccg cgccaagagc cggcggcggc gcatcgcccg 17940 gcggcaccgg agcacccccg ccatgcgcgc ggcgcgagcc ttgctgcgca gggccaggcg 18000 cacgggacgc agggccatgc tcagggcggc cagacgcgcg gcttcaggcg ccagcgccgg 18060 caggacccgg agacgcgcgg ccacggcggc ggcagcggcc atcgccagca tgtcccgcc 18120 gcggcgaggg aacgtgtact gggtgcgcga cgccgccacc ggtgtgcgcg tgcccgtgcg 18180 cacccgcccc cctcgcactt gaagatgttc acttcgcgat gttgatgtgt cccagcggcg 18240 aggaggatgt ccaagcgcaa attcaaggaa gagatgctcc aggtcatcgc gcctgagatc 18300 tacggccctg cggtggtgaa ggaggaaaga aagccccgca aaatcaagcg ggtcaaaaag 18360 gacaaaaagg aagaagaaag tgatgtggac ggattggtgg agtttgtgcg cgagttcgcc 18420 ccccggcggc gcgtgcagtg gcgcgggcgg aaggtgcaac cggtgctgag acccggcacc 18480 accgtggtct tcacgcccgg cgagcgctcc ggcaccgctt ccaagcgctc ctacgacgag 18540 gtgtacgggg atgatgatat tctggagcag gcggccgagc gcctgggcga gtttgcttac 18600 ggcaagcgca gccgttccgc accgaaggaa gaggcggtgt ccatcccgct ggaccacggc 18660 aaccccacgc cgagcctcaa gcccgtgacc ttgcagcagg tgctgccgac cgcggcgccg 18720 cgccgggggt tcaagcgcga gggcgaggat ctgtacccca ccatgcagct gatggtgcc 18780 aagcgccaga agctggaaga cgtgctggag accatgaagg tggacccgga cgtgcagccc 18840 gaggtcaagg tgcggcccat caagcaggtg gccccgggcc tgggcgtgca gaccgtggac 18900 atcaagattc ccacggagcc catggaaacg cagaccgagc ccatgatcaa gcccagcacc 18960 agcaccatgg aggtgcagac ggatccctgg atgccatcgg ctcctagtcg aagaccccgg 19020 cgcaagtacg gcgcggccag cctgctgatg cccaactacg cgctgcatcc ttccatcatc 19080 cccacgccgg gctaccgcgg cacgcgcttc taccgcggtc ataccagcag ccgccgccgc 19140 aagaccacca ctcgccgccg ccgtcgccgc accgccgctg caaccacccc tgccgccctg 19200 gtgcggagag tgtaccgccg cggccgcgca cctctgaccc tgccgcgcgc gcgctaccac 19260 ccgagcatcg ccatttaaac tttcgccagc tttgcagatc aatggccctc acatgccgcc 19320 ttcgcgttcc cattacgggc taccgaggaa gaaaaccgcg ccgtagaagg ctggcgggga 19380 acgggatgcg tcgccaccac caccggcggc ggcgcgccat cagcaagcgg ttggggggag 19440 gcttcctgcc cgcgctgatc cccatcatcg ccgcggcgat cggggcgatc cccggcattg 19500 cttccgtggc ggtgcaggcc tctcagcgcc actgagacac acttggaaac atcttgtaat 19560 aaacccatgg actctgacgc tcctggtcct gtgatgtgtt ttcgtagaca gatggaagac 19620 atcaattttt cgtccctggc tccgcgacac ggcacgcggc cgttcatggg cacctggagc 19680 gacatcggca ccagccaact gaacgggggc gccttcaatt ggagcagtct ctggagcggg 19740 cttaagaatt tcgggtccac gcttaaaacc tatggcagca aggcgtggaa cagcaccaca 19800 gggcaggcgc tgagggataa gctgaaagag cagaacttcc agcagaaggt ggtcgatggg 19860 ctcgcctcgg gcatcaacgg ggtggtggac ctggccaacc aggccgtgca gcggcagatc 19920 aacagccgcc tggacccggt gccgcccgcc ggctccgtgg agatgccgca ggtggaggag 19980 gagctgcctc ccctggacaa gcggggcgag aagcgacccc gccccgatgc ggaggagacg 20040 ctgctgacgc acacggacga gccgcccccg tacgaggagg cggtgaaact gggtctgccc 20100 accacgcggc ccatcgcgcc cctggccacc ggggtgctga aacccgaaaa gcccgcgacc 20160 ctggacttgc ctcctcccca gccttcccgc ccctctacag tggctaagcc cctgccgccg 20220 gtggccgtgg cccgcgcgcg acccgggggc accgcccgcc ctcatgcgaa ctggcagagc 20280 actctgaaca gcatcgtggg tctgggagtg cagagtgtga agcgccgccg ctgctattaa 20340 acctaccgta gcgcttaact tgcttgtctg tgtgtgtatg tattatgtcg ccgccgccgc 20400 tgtccaccag aaggaggagt gaagaggcgc gtcgccgagt tgcaagatgg ccaccccatc 20460 gatgctgccc cagtgggcgt acatgcacat cgccggacag gacgcttcgg agtacctgag 20520 tccgggtctg gtgcagtttg cccgcgccac agacacctac ttcagtctgg ggaacaagtt 20580 taggaacccc acggtggcgc ccacgcacga tgtgaccacc gaccgcagcc agcggctgac 20640 gctgcgcttc gtgcccgtgg accgcgagga caacacctac tcgtacaaag tgcgctacac 20700 gctggccgtg ggcgacaacc gcgtgctgga catggccagc acctactttg acatccgcgg 20760 cgtgctggat cggggcccta gcttcaaacc ctactccggc accgcctaca acagtctggc 20820 ccccaaggga gcacccaaca cttgtcagtg gacatataaa gccgatggtg aaactgccac agaaaaaacc tatacatatg cgtgcagggc attack caaaagatgg tattcaactt ggaactgaca ccgatgatca gccaatctac gcagataaaa cctatcagcc tgaacctcaa gtgggtgatg ctgaatggca tgacatcact ggtactgatg aaaagtatgg aggcagagct cttaagcctg ataccaaaat gaagccttgt tatggttctt ttgccaagcc tactaataa gaggaggtc aggcaaatgt gaaacagga acaggcacta ctaaagaata tgacatagac atggctttct ttgacaacag aagtgcggct gctgctggcc tagctccaga aattgttttg tatactgaaa atgtggattt ggaaactcca throw ttgtatacaa gctcttctat taatttgggt tgcccaacag acctaactac attggtttca gagacaactt tatcgggctc acctactac acagcactgg caatatgggg gtgctggccg gtcaggcttc tcagctgaat gctgtggttg acttgcaaga 21540. cagaacacc gagctgtcct accagctctt gcttgactct ctgggtgaca gaacccggta tttcagtatg tggaatcagg cggtggacag ctatgatcct gatgtgcgca ttattgaaaa 21600 tcatggtgtg gaggatgaac ttcccaacta ttgtttccct ctggatgctg ttggcagaac 21660 agatacttat cagggaatta aggctaatgg aactgatcaa accacatgga ccaaagatga 21720 cagtgtcaat gatgctaatg agataggcaa gggtaatcca ttcgccatgg aaatcaacat 21780 ccaagccaac ctgtggagga acttcctcta cgccaacgtg gccctgtacc tgcccgactc 21840 ttacaagtac acgccggcca atgttaccct gcccaccaac accaacacct acgattacat 21900 gaacggccgg gtggtggcgc cctcgctggt ggactcctac atcaacatcg gggcgcgctg 21960 gtcgctggat cccatggaca acgtgaaccc cttcaaccac caccgcaatg cggggctgcg 22020 ctaccgctcc atgctcctgg gcaacgggcg ctacgtgccc ttccacatcc aggtgcccca 22080 gaaatttttc gccatcaaga gcctcctgct cctgcccggg tcctacacct acgagtggaa 22140 cttccgcaag gacgtcaaca tgatcctgca gagctccctc ggcaacgacc tgcgcacgga 22200 cggggcctcc atctccttca ccagcatcaa cctctacgcc accttcttcc ccatggcgca 22260 caacacggcc tccacgctcg aggccatgct gcgcaacgac accaacgacc agtccttcaa 22320 cgactacctc tcggcggcca acatgctcta ccccatcccg gccaacgcca ccaacgtgcc 22380 catctccatc ccctcgcgca actgggccgc cttccgcggc tggtccttca cgcgtctcaa 22440 gaccaaggag acgccctcgc tgggctccgg gttcgacccc tacttcgtct actcgggctc 22500 catcccctac ctcgacggca ccttctacct caaccacacc ttcaagaagg tctccatcac 22560 cttcgactcc tccgtcagct ggcccggcaa cgaccggctc ctgacgccca acgagttcga 22620 aatcaagcgc accgtcgacg gcgagggcta caacgtggcc cagtgcaaca tgaccaagga 22680 ctggttcctg gtccagatgc tggcccacta caacatcggc taccagggct tctacgtgcc 22740 cgagggctac aaggaccgca tgtactcctt cttccgcaac ttccagccca tgagccgcca 22800 ggtggtggac gaggtcaact acaaggacta ccaggccgtc accctggcct accagcacaa 22860 caactcgggc ttcgtcggct acctcgcgcc caccatgcgc cagggccagc cctaccccgc 22920 caactacccc tacccgctca tcggcaagag cgccgtcacc agcgtcaccc agaaaaagtt 22980 cctctgcgac agggtcatgt ggcgcatccc cttctccagc aacttcatgt ccatgggcgc 23040 gctcaccgac ctcggccaga acatgctcta tgccaactcc gcccacgcgc tagacatgaa 23100 tttcgaagtc gaccccatgg atgagtccac ccttctctat gttgtcttcg aagtcttcga 23160 cgtcgtccga gtgcaccagc cccaccgcgg cgtcatcgag gccgtctacc tgcgcacccc 23220 cttctcggcc ggtaacgcca ccacctaagc tcttgcttct tgcaagccat ggccgcgggc 23280 tccggcgagc aggagctcag ggccatcatc cgcgacctgg gctgcgggcc ctacttcctg 23340 ggcaccttcg ataagcgctt cccgggattc atggccccgc acaagctggc ctgcgccatc 23400 gtcaacacgg ccggccgcga gaccgggggc gagcactggc tggccttcgc ctggaacccg 23460 cgctcgaaca cctgctacct cttcgacccc ttcgggttct cggacgagcg cctcaagcag 23520 atctaccagt tcgagtacga gggcctgctg cgccgcagcg ccctggccac cgaggaccgc 23580 tgcgtcaccc tggaaaagtc cacccagacc gtgcagggtc cgcgctcggc cgcctgcggg 23640 ctcttctgct gcatgttcct gcacgccttc gtgcactggc ccgaccgccc catggacag23700 aaccccacca tgaacttgct gacggggggtg cccaacggca tgctccagtc gccccaggtg 23820. 23820. 23820. 23820. 23820. 23820. 23820. 23820 tactttcgct cccaccgcgc gcgcatcgag aaggccaccg ccttcgaccg catgaatcaa 23940. ccgtgtgtgt atgttaaatg tctttaataa acagcacttt catgttacac atgcatctga gatgattt ttagaaatcg aaagggttct gccgggtctc ggcatggccc gcgggcaggg acacgttgcg gaactggtac ttggccagcc acttgaactc ggggatcagc agtttgggca gcggggtgtc ggggaaggag tcggtccaca gcttccgcgt cagttgcagg 24120. gcgcccagca ggtcgggcgc ggagatcttg aaatcgcagt tgggacccgc gttctgcgcg 24180. 24240. cgggagttgc ggtacacggg gttgcagcac tggacacca tcagggccgg gtgcttcacg ctcgccagca ccgtcgcgtc ggtgatgctc tccacgtcga ggtcctcggc gttggccatc 24300 ccgaaggggg tcatcttgca ggtctgcctt cccatggtgg gcacgcaccc gggcttgtgg 24360 ttgcaatcgc agtgcagggg gatcagcatc atctgggcct ggtcggcgtt catccccggg 24420 tacatggcct tcatgaaagc ctccaattgc ctgaacgcct gctgggcctt ggctccctcg 24480 gtgaagaaga ccccgcagga cttgctagag aactggttgg tggcgcaccc ggcgtcgtgc 24540 acgcagcagc gcgcgtcgtt gttggccagc tgcaccacgc tgcgccccca gcggttctgg 24600 gtgatcttgg cccggtcggg gttctccttc agcgcgcgct gcccgttctc gctcgccaca 24660 tccatctcga tcatgtgctc cttctggatc atggtggtcc cgtgcaggca ccgcagcttg 24720 ccctcggcct cggtgcaccc gtgcagccac agcgcgcacc cggtgcactc ccagttcttg 24780 tgggcgatct gggaatgcgc gtgcacgaag ccctgcagga agcggcccat catggtggtc 24840 agggtcttgt tgctagtgaa ggtcagcgga atgccgcggt gctcctcgtt gatgtacagg 24900 tggcagatgc ggcggtacac ctcgccctgc tcgggcatca gctggaagtt ggctttcagg 24960 tcggtctcca cgcggtagcg gtccatcagc atagtcatga tttccatacc cttctcccag 25020 gccgagacga tgggcaggct catagggttc ttcaccatca tcttagcgct agcagccgcg 25080 gccagggggt cgctctcgtc cagggtctca aagctccgct tgccgtcctt ctcggtgatc 25140 cgcaccgggg ggtagctgaa gcccacggcc gccagctcct cctcggcctg tctttcgtcc 25200 tcgctgtcct ggctgacgtc ctgcaggacc acatgcttgg tcttgcgggg tttcttcttg 25260 ggcggcagcg gcggcggaga tgttggagat ggcgaggggg agcgcgagtt ctcgctcacc 25320 actactatct cttcctcttc ttggtccgag gccacgcggc ggtaggtatg tctcttcggg 25380 ggcagaggcg gaggcgacgg gctctcgccg ccgcgacttg gcggatggct ggcagagccc 25440 cttccgcgtt cgggggtgcg ctcccggcgg cgctctgact gacttcctcc gcggccggcc 25500 attgtgttct cctagggagg aacaacaagc atggagactc agccatcgcc aacctcgcca 25560 tctgccccca ccgccgacga gaagcagcag cagcagaatg aaagcttaac cgccccgccg 25620 cccagccccg ccacctccga cgcggccgtc ccagacatgc aagagatgga ggaatccatc 25680 gagattgacc tgggctatgt gacgcccgcg gagcacgagg aggagctggc agtgcgcttt 25740 tcacaagaag agatacacca agaacagcca gagcagagaag cagagaatga cgagagtcag 25800 gctgggctcg agcatgacgg cgactacctc cacctgagcg ggggggagga cgcgctcatc 25860 aagcatctgg cccggcaggc caccatcgtc aagcattgcgc tgctcgaccg caccgaggtg 25920 cccctcagcg tggaggagct cagccgcgcc tacgagttga acctctttc gccgcgcgtg 25980 ccccccaagc gccagcccaa tggcacctgc gagcccaacc cgcgcctcaa cttctacccg 26040 gtttcgcgg tgcccgaggc cctggccacc taccacatct ttttcaagaa ccaaaagatc 26100 cccgtctcct gccgcgccaa ccgcacccgc gccgacgcccc ttttcaacct gggtcccggc 26160 gcccgcctac ctgatatcgc ctccttggaa gaggttccca agatcttcga gggtctgggc 26220 agcgacgaga ctcgggccgc gaacgctctg caaggaaag gaggagagca tgagcaccac 26280 agcgccctgg tcgagttgga agggcaac gcgcggctgg cggtgctcaa acgcacggtc 26340 gagctgaccc atttcgccta cccggctctg aacctgcccc ccaaagtcat gagcgcggtc 26400 atggaccagg tgctcatcaa gcgcgcgtcg cccatctccg aggacgaggg catgcaagac 26460 tccgaggagg gcaagcccgt ggtcagcgac gagcagctgg cccggtggct gggtcctaat 26520 gctagtcccc agagtttgga agagcggcgc aaactcatga tggccgtggt cctggtgacc 26580 gtggagctgg agtgcctgcg ccgcttcttc gccgacgcgg agaccctgcg caaggtcgag 26640 gagaacctgc actacctctt caggcacggg ttcgtgcgcc aggcctgcaa gatctccaac 26700 gtggagctga ccaacctggt ctcctacatg ggcatcttgc acgagaaccg cctggggcag 26760 aacgtgctgc acaccaccct gcgcggggag gcccggcgcg actacatccg cgactgcgtc 26820 tacctctacc tctgccacac ctggcagacg ggcatgggcg tgtggcagca gtgtctggag 26880 gagcagaacc tgaaagagct ctgcaagctc ctgcagaaga acctcaaggg tctgtggacc 26940 gggttcgacg agcgcaccac cgcctcggac ctggccgacc tcattttccc cgagcgcctc 27000 aggctgacgc tgcgcaacgg cctgcccgac tttatgagcc aaagcatgtt gcaaaacttt 27060 cgctctttca tcctcgaacg ctccggaatc ctgcccgcca cctgctccgc gctgccctcg 27120 gacttcgtgc cgctgacctt ccgcgagtgc cccccgccgc tgtggagcca ctgctacctg 27180 ctgcgcctgg ccaactacct ggcctaccac tcggacgtga tcgaggacgt cagcggcgag 27240 ggcctgctcg agtgccactg ccgctgcaac ctctgcacgc cgcaccgctc cctggcctgc 27300 aacccccagc tgctgagcga gacccagatc atcggcacct tcgagttgca agggcccagc 27360 gaaggcgagg gttcagccgc caaggggggt ctgaaactca ccccggggct gtggacctcg 27420 gcctacttgc gcaagttcgt gcccgaggac taccatccct tcgagatcag gttctacgag 27480 gaccaatccc atccgcccaa ggccgagctg tcggcctgcg tcatcaccca gggggcgatc 27540 ctggcccaat tgcaagccat ccagaaatcc cgccaagaat tcttgctgaa aaagggccgc 27600 ggggtctacc tcgaccccca gaccggtgag gagctcaacc ccggcttccc ccaggatgcc 27660 ccgaggaaac aagaagctga aagtggagct gccgcccgtg gaggatttgg aggaagactg 27720 ggagaacagc agtcaggcag aggagga gatgaggaa gactgggaca gcactcaggc 27780 agaggaggac agcctgcaag acagtgga ggagacgag gaggaggcag agggaggt 27840 ggaagaagca gccgccgcca gaccgtcgtc ctcggcgggg gagaagcaa gcagcacgga 27900 taccatctcc gctccggggtc ggggtcccgc tcgaccac agtagatgggg acgagaccgg 27960 acgattcccg aaccccacca cccagaccgg tagaggag cggcagggat acagtccctg 28020 gcgggggcac aaaaacgcca tcgtctcctg cttgcaggcc tgcgggggca acatctcctt 28080 cacccggcgc tacctgctt tccaccgcgg ggtgaactt cccgcaaca tcttgcatta 28140 ctaccgtcac ctccacagcc cctactactt ccagagag gcagcagcag cagaaaaga 28200 ccagcagaaa accagcagct agaaatcca cagcggcggc agcaggtgga ctgaggatcg 28260 cggcgaacga gccggcgcaa acccgggagc tgaggaccg gatctttccc acccttatg 28320 ccatcttcca gcagagtcgg gggcaggagc aggactgaa agtcagaac cgttctctgc 28380 gctcgctcac ccgcagttgt ctgtatcaca agagcgaaga ccaacttcag cgcactctcg 28440 aggacgccga ggctctcttc aacaagtact gcgcgctcac tcttaaagag tagcccgcgc 28500 ccgcccagtc gcagaaaaag gcgggaatta cgtcacctgt gcccttcgcc ctagccgcct 28560 ccacccatca tcatgagcaa agagattccc acgccttaca tgtggagcta ccagccccag 28620 atgggcctgg ccgccggtgc cgcccaggac tactccaccc gcatgaattg gctcagcgcc 28680 gggcccgcga tgatctcacg ggtgaatgac atccgcgccc accgaaacca gatactccta 28740 gaacagtcag cgctcaccgc cacgccccgc aatcacctca atccgcgtaa ttggcccgcc 28800 gccctggtgt accaggaaat tccccagccc acgaccgtac tacttccgcg agacgcccag 28860 gccgaagtcc agctgactaa ctcaggtgtc cagctggcgg gcggcgccac cctgtgtcgt 28920 caccgccccg ctcagggtat aaagcggctg gtgatccggg gcagaggcac acagctcaac 28980 gacgaggtgg tgagctcttc gctgggtctg cgacctgacg gagtcttcca actcgccgga 29040 tcggggagat cttccttcac gcctcgtcag gccgtcctga ctttggagag ttcgtcctcg 29100 cagccccgct cgggtggcat cggcactctc cagttcgtgg aggagttcac tccctcggtc 29160 tacttcaacc ccttctccgg ctcccccggc cactacccgg acgagttcat cccgaacttc 29220 gacgccatca gcgagtcggt ggacggctac gattgaatgt cccatgtcga cccccggtcc 29280 cccacccagt cccccgagga ggtccgcaaa tgcaaattcc aagaaccctg gaaattcctc 29340 aaatgctacc gccaaaaatc agacatgcat cccagctgga tcatgatcat tgggatcgtg 29400 aacattctgg cctgcaccct catctcctttt gtgatttacc cctgctttga cttttggttgg 29460 aactcgccag aggcgcta tctcccgcct gaacctgaca caccaccaca gcaacctcag 29520 gcacacgcac taccaccact acagcctagg ccacaataca tgcccatatt agactatgag 29580 gccgagccac agcgacccat gctccccgct attagttact tcaatctaac cggcggagat 29640 gactgaccca ctggccaaca acaacgtcaa cgaccttctc ctggacatgg acggccgcgc 29700 ctcggagcag cgactcgccc aacttcgcat tcgccagcag caggagagag ccgtcaagga 29760 gctgcaggat gcggtggcca tccaccagtg caagagaggc atcttctgcc tggtgaaaca 29820 ggccaagatc tcctacgagg tcactccaaa cgaccatcgc ctctcctacg agctcctgca 29880 gcagcgccag aagttcacct gcctggtcgg agtcaaccc atcgtcatca cccagcagtc 29940 tggcgatacc aaggggtgca tccactgctc ctgcgactcc cccgactgcg tccacactct 30060. gatcaagacc ctctgcggcc tccgcgacct cctccccatg aactaatcac ccccttatcc agtgaata agatcatatt agtgaatt ttacagaat aaaaaatat catttgattt gaataaga tacaatcata ttgatgattt gagtttaaca aaaaaataa gathering cttgaatct gataccaggt ctctgtccat gttttctgcc aacaccactt cactcccctc ttcccagctc tggtactgca ggccccggcg ggctgcaaac ttcctccaca cgctgaaggg 30300. 30360. gatgtcaaat tcctcctgtc cctcaatctt cattttatct tctatcagat gtccaaaaag cgcgtccggg tggatga cttcgacccc gtctacccct acgatgcaga caacgcaccg accgtgccct tcatcaaccc ccccttcgtc tcttcagatg gattccaaga gaagcccctg ggggtgttgt ccctgcgact ggccgacccc gtcaccacca agaacgggga aatcaccctc 30540 aagctgggag agggggtgga cctcgattcc tcgggaaaac tcatctccaa cacggccacc aaggccgccg cccctctcag tttttccaac aacaccattt cccttaacat ggatcacccc ttttacacta aagatggaa attatcctta caagtttctc cracked tatactgaga acaagcattc taaacacact agctttaggt tttggatcag gtttaggact ccgtggctct gccttggcag tacagttagt ctctccactt acatttgata ctgatggaaa cataaagctt accttagaca gaggtttgca tgttacaaca ggagatgca ttgaaagca cataagctgg gctaaaggtt taaaatttga agatggagcc atagcaacca acattggaa tgggttagag tttggaagca gtagtacaga aacaggtgtt gatgatgctt acccaatcca agttaaactt ggatctggcc ttagctttga cagtacagga gccataatgg ctggtaacaa agaagacgat aaactcactt tgtggacaac acctgatcca tcaccaaact gtcaaatact cgcagaaaat gatgcaaaac taacactttg cttgactaaa tgtggtagtc aaatactggc cactgtgtca gtcttagttg taggagtgg aacctaaac cccattactg gcaccgtaag cagtgctcag 31260 gtgtttctac gtttgatgc aaacggtgtt cttttacag aacattctac actaaaaaaa 31320 tactgggggt ataggcagggg agatagcata gatggcactc catataccaa tgctgtagga 31380 ttcatgcccca atttaaaagc ttcacaag tcacaagtt ctactactaa aaatatata 31440 gtagggcaag tatacatgaa tggagatt tcaaaccta tgctctcac tataaccctc 31500 aatggtactg atgacagca cagtacatat tcaatgtcat ttcatacac ctggactaat 31560 ggaagctatg ttggagcaac atttggggct aactcttata ccttctcata catcgcccaa 31620 gatgaacac tgtatcccac cctgcatgcc aacccttccc accccactct gtggacaaaa 31680 ctctgaaca CAAAAAAA taagttca gtgttttatt gattcacag ttttacagga 31740 ttcgagcagt tattttcct ccacccccc aggacatgga atacaccacc ctctcccccc 31800 gcacagcctt gaacatctga atgccattgg tgatggacat gctttggtc tccacgttcc 31860 acacagtttc agagcgagcc agtctcgggt cggtcaggga gatgaaaccc tccgggcact 31920 cccgcatctg cacctcacag ctcaacagct gaggattgtc ctcggtggtc gggatcacgg 31980 ttatctggaa gaagcagaag agcggcggtg ggaatcatag tccgcgaacg ggatcggccg 32040 gtggtgtcgc atcaggcccc gcagcagtcg ctgccgccgc cgctccgtca agctgctgct 32100 cagggggtcc gggtccaggg actccctcag catgatgccc acggccctca gcatcagtcg 32160 tctggtgcgg cgggcgcagc agcgcatgcg gatctcgctc aggtcgctgc agtacgtgca 32220 acacagaacc accaggttgt tcaacagtcc atagttcaac acgctccagc cgaaactcat 32280 cgcgggaagg atgctaccca cgtggccgtc gtaccagatc ctcaggtaaa tcaagtggtg 32340 ccccctccag aacacgctgc ccacgtacat gatctccttg ggcatgtggc ggttcaccac 32400 ctcccggtac cacatcaccc tctggttgaa catgcagccc cggatgatcc tgcggaacca 32460 cagggccagc accgccccgc ccgccatgca gcgaagagac cccgggtccc ggcaatggca 32520 atggaggacc caccgctcgt acccgtggat catctgggag ctgaacaagt ctatgttggc 32580 acagcacagg catatgctca tgcatctctt cagcactctc aactcctcgg gggtcaaaac 32640 catatcccag ggcacgggga actcttgcag gacagcgaac cccgcagaac agggcaatcc 32700 tcgcacagaa cttacattgt gcatggacag ggtatcgcaa tcaggcagca ccgggtgatc 32760 ctccaccaga gaagcgcggg tctcggtctc ctcacagcgt ggtaaggggg ccggccgata 32820 cgggtgatgg cgggacgcgg ctgatcgtgt tcgcgaccgt gtcatgatgc agttgctttc 32880 ggacattttc gtacttgctg tagcagaacc tggtccgggc gctgcacacc gatcgccggc 32940 ggcggtctcg gcgcttggaa cgctcggtgt tgaaattgta aaacagccac tctctcagac 33000 cgtgcagcag atctagggcc tcaggagtga tgaagatccc atcatgcctg atggctctga 33060 tcacatcgac caccgtggaa tgggccagac ccagccagat gatgcaattt tgttgggttt 33120 cggtgacggc gggggaggga agaacaggaa gaaccatgat taacttttaa tccaaacggt 33180 ctcggagtac ttcaaaatga agatcgcgga gatggcacct ctcgcccccg ctgtgttggt 33240 ggaaaataac agccaggtca aaggtgatac ggttctcgag atgttccacg gtggcttcca 33300 gcaaagccctc cacgcgcaca tccagaaca agacaatagc gaagcggga gggttctctta 33360 attcctcaat catcatgtta cactcctgca ccatccccag atatttca ttttccagc 33420 cttgaatgat tcgaactagt tcgtgaggta atccaagcc agccatgata aagagctcgc 33480 gcagagcgcc ctccaccggc attcttaagc acaccctcat attccaaga tattctgctc 33540 ctggttcacc tgcagcagat tgacaagcgg atatcaaa tctctgccgc gatccctgag 33600 ctcctccctc agcaatact gtaagtactc ttcatatcc tctccgaat tttagccat 33660 aggaccacca ggaataagat taggcaagc cacagtacag ataaccgaa gtcctcccca 33720 gtgagcattg ccaatgcaa gactgctata agcatgctgg ctagacccgg tgatatctctc 33780 cagataactg gagaaaat cgcccaggca atttttaag aaatcaaaaaaatc 33840 ctccaggtgg acgtttagag cctcgggaac acgatgaag taatgcaag cggtgcgttc 33900 cagcatggtt agttagctga tctgtagaa aaaaaaaat gaacattaaa ccatgctagc 33960 ctggcgaaca ggtgggtaaa tcgttctctc cagcaccagg caggccacgg ggtctccggc 34020 gcgaccctcg taaaaattgt cgctatgatt gaaaaccacc acagagac gttcccggtg 34080 gccggcgtga atgattcgac aagatgata cacccccgga acatggcgt ccgcgagtga 34140 aaaaaagcgc ccgaggaagc aaaggcac tacaatgctc agtaaataa tctcaagctcc 34200 agcaaagcga tgccatgcgg atgaagcaca aaattctcag gtgcgtacaa atgtaatta 34260 ctcccctcct gcacaggcag caagcccc gatccctcca ggtacacata caagcctca 34320 gcgtccatag cttaccgagc agcagcacac aacaggcgca agatcagag aaggctgag 34380 ctctaacctg tccacccgct ctctgctcaa tatatagccc agatctacc tgacgtaag 34440 gccaaagtct aaaaatacccc gccaaataat cacacacgcc cagcacacgc caccaaccg 34500 gtgacacact caaaaaata cgcgcactc ctcaacgcc caaactgcc gtcattccg 34560 ggttcccacg ctacgtcatc aaaaacacgac ttcaattc cgtcgaccgt taaaaacgtc 34620 acccgccccg cccctaacgg tcgcccgtct ctcagccaat cagcgccccg catccccaaa 34680 ttcaaacacc tcatttgcat attaacgcgc acaaaaagtt tgaggtatat tattgatgat 34740 gg 34742

Claims

1. A pharmaceutical preparation for inducing specific immunity against SARS-CoV-2, the pharmaceutical preparation comprising component 1, said component 1 comprising an expression vector based on the genome of recombinant human adenovirus serotype 26, wherein the E1 and E3 regions are deleted from said genome and the ORF6-Ad26 region is replaced by ORF6-Ad5, wherein an expression cassette selected from SEQ ID NO:1, SEQ ID NO:2, and SEQ ID NO:3 is disposed therein, and said pharmaceutical preparation further comprising component 2, said component 2 comprising an expression vector based on the genome of recombinant simian adenovirus serotype 25, wherein the E1 and E3 regions are deleted from said genome, wherein an expression cassette selected from SEQ ID NO:4, SEQ ID NO:2, and SEQ ID NO:3 is disposed therein.

2. The pharmaceutical preparation according to claim 1, characterized in that, The pharmaceutical preparation is formulated as a liquid preparation or a lyophilized preparation.

3. The pharmaceutical preparation according to claim 2, characterized in that, Buffer solutions used in liquid formulations contain, by mass percentage:

4. The pharmaceutical preparation according to claim 2, characterized in that, The buffer solution used for lyophilized formulations comprises, by mass percentage:

5. The pharmaceutical preparation according to claim 1, characterized in that, Component 1 and Component 2 are placed in different packages.

6. A pharmaceutical preparation for inducing specific immunity against SARS-CoV-2, the pharmaceutical preparation comprising component 1, said component 1 comprising an expression vector based on the genome of recombinant simian adenovirus serotype 25, wherein E1 and E3 regions are deleted from said genome, and wherein an expression cassette selected from SEQ ID NO:4, SEQ ID NO:2, and SEQ ID NO:3 is placed therein, and said pharmaceutical preparation further comprising component 2, said component 2 comprising an expression vector based on the genome of recombinant human adenovirus serotype 5, wherein E1 and E3 regions are deleted from said genome, and wherein an expression cassette selected from SEQ ID NO:1, SEQ ID NO:2, and SEQ ID NO:3 is placed therein.

7. The pharmaceutical preparation according to claim 6, characterized in that, The pharmaceutical preparation is formulated as a liquid preparation or a lyophilized preparation.

8. The pharmaceutical preparation according to claim 7, characterized in that, Buffer solutions used in liquid formulations contain, by mass percentage:

9. The pharmaceutical preparation according to claim 7, characterized in that, The buffer solution used for lyophilized formulations comprises, by mass percentage:

10. The pharmaceutical preparation according to claim 6, characterized in that, Component 1 and Component 2 are placed in different packages.

11. Use of the pharmaceutical formulation of claim 1 or 6 in the preparation of a medicament for inducing specific immunity against SARS-CoV-2, wherein components 1 and 2 are used in effective amounts sequentially, with an interval of not less than one week.

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