Compositions and methods for mrs vaccines against novel coronavirus infection

Abstract

Compositions and methods are provided for effective mRNA vaccines for the prevention and treatment of SARS-CoV-2 infection. These compositions include a pharmaceutical composition containing one or more mRNA molecules encoding epitopes of the spike protein, including mutated epitopes, or a cocktail of mRNAs encoding key viral genes, along with pharmaceutically acceptable polymeric nanoparticle carriers and liposomal nanoparticle carriers. Methods for stimulating systemic immune responses and treatment are provided, including subcutaneous injection, intraperitoneal injection, and intramuscular injection.

Description

[0001] This application claims priority to U.S. Provisional Application No. 62 / 971,834, filed February 7, 2020; No. 63 / 058,463, filed July 29, 2020; and No. 63 / 130,581, filed December 24, 2020, the contents of which are incorporated herein by reference in their entirety. Technical Field

[0002] It provides information on vaccination, prevention, and treatment of SARS-CoV-2 infection, as well as methods of administration. Background Technology

[0003] Novel Coronavirus (SARS-CoV-2) Infection: Biology and Pathology Coronaviruses (CoVs) have crossed species barriers multiple times, and some have become important human pathogens. In the past 20 years, two coronaviruses that infect animals have evolved and caused outbreaks in humans: SARS-CoV (2002, genus *Betacoronavirus*, subgenus *Sarbecovirus*) and MERS-CoV (2012, genus *Betacoronavirus*, subgenus *Merbecovirus*) [Drosten et al., New Engl J Med. 2003; 348: 1967-1976; Zaki et al., New Engl J Med. 2012; 367: 1814-1820]. SARS-CoV-2 appears to be transmitted through respiratory droplets in person-to-person contact [Li et al., NEngl J Med. 2020; 382:1199-1207].

[0004] Genetic analysis revealed that SARS-CoV-2 is closely related to SARS-CoV and genetically clusters within the genus *Betacoronavirus*, forming a distinct lineage of the subgenus *Sabecoronavirus* B along with two bat-derived SARS-like strains. The viral genome consists of six major open reading frames (ORFs) common to coronaviruses, along with several accessory genes. Further analysis showed that some sequences in the SARS-CoV-2 genome share less than 80% with SARS-CoV. However, the seven conserved replicasease (RdRP) domains in the ORF1ab, used for CoV species classification, share 94.6% AA sequence similarity between SARS-CoV-2 and SARS-CoV, suggesting they belong to the same species. Angiotensin-converting enzyme II (ACE2) is an extracellular peptidase known as the cellular receptor for SARS-CoV. This receptor has also been shown to be used by SARS-CoV-2 to enter human cells. Domains 266-330 (based on the sequence number of the mature protein) have been shown to be essential for the interaction between SARS-CoV-2 and ACE2 [Veljkovic et al., F1000Research. 2020; 9: 52.].

[0005] Current prevention and treatment of the novel coronavirus (SARS-CoV-2) Currently, the Chinese CDC and other organizations have submitted 287,543 complete genome sequences of SARS-CoV-2 to GISAID [GISAID, Newly discovered betacoronavirus, Wuhan 2019-2020 (2019); https: / / www.gisaid.org / ]. One of these sequences has been published in GenBank (Accession: MN908947.3) [http: / / www.ncbi.nlm.nih.gov / genbank / ]. Currently, there are only two vaccines with emergency use authorization, and no licensed vaccines for the prevention of coronavirus infections, including SARS-CoV-2 infection.

[0006] Advantages of mRNA vaccines Vaccination is the most successful medical approach to prevent and control disease. The successful development and use of vaccines has saved thousands of lives and saved a great deal of money. A key advantage of RNA vaccines is that RNA can be produced in the laboratory from DNA templates using readily available materials, which is cheaper and faster than traditional vaccine production, which may require the use of chicken eggs or other mammalian cells. In addition, mRNA vaccines have the potential to simplify vaccine discovery and development and facilitate a rapid response to emerging infectious diseases [Maruggi et al., Mol Ther. 2019; 27(4):757-772].

[0007] Preclinical and clinical trials have demonstrated that mRNA vaccines provide safe and durable immune responses in animal models and humans. mRNA vaccines against infectious diseases can be developed as preventative or therapeutic agents. mRNA vaccines expressing antigens of infectious pathogens have been shown to induce effective T-cell and humoral immune responses [Pardi et al., Nat Rev Drug Discov. 2018; 17: 261–279.]. Compared to the production of whole microorganisms, live attenuated vaccines, and subunit vaccines, the production process for mRNA vaccines is cell-free, simple, and rapid. This rapid and simple production process makes mRNA a promising bioproduct that has the potential to bridge the gap between emerging infectious diseases and the urgent need for effective vaccines. Summary of the Invention

[0008] A composition is provided comprising an effective amount of messenger ribonucleic acid (mRNA) formulated in a pharmaceutically acceptable carrier, the messenger ribonucleic acid containing an open reading frame (ORF) encoding a SARS-CoV-2 protein or protein fragment, wherein the mRNA and the carrier form polymeric nanoparticles. The SARS-CoV-2 protein may be a spike (S) protein or a spike (S) subunit. The ORF may be mRNA encoding a peptide, such as a peptide having a sequence selected from SEQ ID Nos. 14 to 31 or 50-66. The ORF may comprise, for example, mRNA having a sequence shown in SEQ ID Nos. 32 to 49 or SEQ ID Nos. 91 to 107. The ORF may be selected from mRNA molecules encoding amino acid sequences of SEQ ID Nos. 11, 13, 68, 70, 72, 74, and 90. The ORF may have a sequence such as SEQ ID Nos. 1-4, 10, 12, 67, 69, 71, 73, or 89.

[0009] These compositions may also contain all or a portion of mRNA containing orf1a and / or orf1b, wherein the mRNA encodes SARS-CoV-2 nonstructural protein polymerase and / or RdRp. The ORF may be an mRNA having, for example, a sequence of SEQ ID NO: 5 or 6. The ORF may encode all or a portion of SARS-CoV-2, encoding envelope (E), membrane (M), and / or nucleocapsid (N) proteins. These compositions may also contain one or more mRNA sequences encoding all or a portion of SARS-CoV-2, encoding envelope (E), membrane (M), and / or nucleocapsid (N) proteins. The ORF may be selected from mRNA molecules having sequences as shown in SEQ ID No. 7, 8, or 9.

[0010] In any of these compositions, the mRNA may be chemically modified with one or more N1-methylpseudouridine and / or pseudouridine residues. The uridine residues in the mRNA may be all N1-methylpseudouridine or pseudouridine.

[0011] In any of these compositions, the mRNA molecule may further comprise a 3'-UTR, a 5'-UTR, and optionally further comprise additional elements that (a) stabilize the composition and (b) enhance the expression of the polypeptide encoded by the ORF. The 5'-UTR may contain an m7G cap structure and a start codon, and / or the 3'-UTR may contain a stop codon and a polyA tail.

[0012] The composition may contain more than one ORF and may encode at least two polypeptides.

[0013] The composition may contain a polymer nanoparticle carrier containing a histidine-lysine copolymer (HKP). Suitable examples of HKP include, but are not limited to, polymers having side chains selected from SEQ ID NO: 75-88.

[0014] A method for preparing the above composition by expressing the ORF in a linear in vitro transcription (IVT) system or by a plasmid DNA (pDNA) vector delivery system is also provided.

[0015] The above composition can be prepared by contacting an ORF-containing composition with HKP, wherein the mRNA and HKP self-assemble into nanoparticles. The composition may further contain lipids, wherein the lipids may optionally be cationic lipids.

[0016] A method for treating a subject with SARS-CoV-2 infection by administering a pharmaceutically effective amount of the above composition to the subject is also provided. The composition can be nebulized via a nebulizer inhalation system before or during administration. The nebulizer system can be a portable nebulizer for whole-respiratory drug delivery. The composition can be administered by subcutaneous injection, intramuscular injection, or intraperitoneal injection (ip). Attached Figure Description

[0017] Figure 1 A schematic diagram of an optimized mRNA vaccine expression structure is shown. This structure is contained in a linear in vitro transcription (IVT) expression system or a plasmid DNA delivery vector.

[0018] Figure 2 The results of immunofluorescence analysis of spike protein expression after transient mRNA transfection are shown. Transfection conditions: ThermoFisher Scientific Lipofectamine MessengerMAX transfection reagent protocol (Pin. No. MAN0010803 Rev D.0).

[0019] Figure 3 The results of Western blot analysis of spike protein expression after transient mRNA transfection are shown. Transfection conditions: ThermoFisher Scientific Lipofectamine MessengerMAX transfection reagent protocol (Pin. No. MAN0010803 Rev D.0). Primary antibody: SARS-CoV / SARS-CoV-2 spike protein S2 monoclonal antibody (ThermoFisher Scientific, MA5-35946). Secondary antibody: Mouse IgG (H+L) secondary antibody (ThermoFisher Scientific, A24512). Internal control: HRP anti-β Actin antibody (abcam, ab49900).

[0020] Figure 4 The spike protein ORF mRNA sequence is shown.

[0021] Figure 5 The spike protein ORF mRNA and protein sequence of the D614G mutation are shown.

[0022] Figure 6 The SARS RBD mRNA sequence and the amino acid sequence of the D614G mutated spike protein ORF are shown.

[0023] Figure 7The mRNA and amino acid sequences of the polypeptide epitopes for SARS-CoV-2 are shown.

[0024] Figure 8 The D614G_S6P DNA and amino acid sequence are shown.

[0025] Figure 9 The RBD trimer DNA and amino acid sequence are shown.

[0026] Figure 10 The DNA and amino acid sequence of VUI-202012 are shown.

[0027] Figure 11 The HKP peptide sequence is shown.

[0028] Figure 12 The D614G S2P DNA sequence is shown.

[0029] Figure 13 The amino acid sequence of D614G S2P is shown.

[0030] Figure 14 The sequence of the mRNA molecule encoding the peptide epitope of SARS-CoV-2 is shown. Detailed Implementation

[0031] Prophylactic and therapeutic agents for vaccination, prevention, and treatment of SARS-CoV-2 infection are provided. Pharmaceutical compositions containing mRNA sequences formulated with pharmaceutical excipients are provided. The mRNA-based pharmaceutical formulations contain mRNA sequences encoding viral proteins essential for coronavirus infection and replication, along with a specific vector. Suitable vectors include histidine-lysine copolymers (HKPs) and / or HKPs plus lipids (e.g., cationic lipids). Methods of using these pharmaceutical compositions are also provided, including therapeutic methods, process development, and specific delivery routes and protocols.

[0032] Target selection and design of mRNA vaccines against SARS-CoV-2 At the protein level, predicted genes for SARS-CoV-2 were identified and annotated. The results showed that the annotations for all six SARS-CoV-2 genomes were identical, including seven functional proteins validated in SARS-CoV-like coronaviruses (pp1a, E2 glycoprotein precursor, hypothetical protein sars3a, matrix protein, hypothetical protein sars6, hypothetical protein sars7a, and nucleocapsid protein) [B. Buchfink, et al., Nat Methods. 2015; 12: 59-60]. These results illustrate the similarities and differences between SARS-CoV-2 and SARS-CoV or MERS-CoV-like coronaviruses.

[0033] The primary human receptor for SARS-CoV-2 is believed to be human angiotensin-converting enzyme 2 (ACE2). Domains 288-330 of the S1 protein from SARS-CoV-2 represent promising therapeutic and / or vaccine targets [Veljkovi et al., ibid.]. Generally, the spike (S) protein of coronaviruses including SARS-CoV-2 is a major inducer of neutralizing antibodies, and the receptor-binding domain (RBD) in the S1 subunit of the S protein contains multiple conformational neutralizing epitopes [He et al., JImmunol.2005; 174(8): 4908-4915]. This suggests that recombinant proteins containing the RBD and vectors including mRNA encoding the RBD sequence could be used to develop safe and effective SARS-CoV-2 vaccines.

[0034] As described herein, a novel mRNA vaccine (mRNA-1273) has been developed using an immunogenic composition containing messenger ribonucleic acid (mRNA) with an open reading frame (ORF) encoding one or more epitopes of one or more proteins derived from SARS-CoV-2. The mRNA is formulated in a pharmaceutically acceptable carrier containing polymeric nanoparticles and / or liposome nanoparticles. The composition can be administered to the subject in an amount sufficient to effectively induce a specific immune response against SARS-CoV-2. The composition may contain more than one mRNA molecule. Each additional copy enhances the immunostimulatory activity of the composition. All mRNAs encode antigens that are highly immunogenic and essential for SARS-CoV-2 entry into epithelial cells (the first step in SARS-CoV-2 infection).

[0035] In one embodiment, the composition contains one or more mRNAs encoding the entire spike (S) protein of SARS-CoV-2. Figure 3The S protein is used by the virus to attach to host cell receptors. The mRNA sequence encoding the spike protein is very long; therefore, to improve the efficiency of mRNA expression system preparation, it is divided into four regions, one, some, or all of which can be used as mRNA antigens targeting the spike protein structure when expressed within the framework of the spike protein domain (SEQ ID No. 1 to 4). Twenty-four short peptide fragments as potential epitopes were selected as targets (SEQ ID No. 14 to 31). The selected short peptides are less than 20 amino acids, advantageously less than 15 amino acids. The mRNA sequences encoding the short peptide sequences were designed using methods well-known in the art (SEQ ID No. 32 to 49).

[0036] Furthermore, compared to other coronaviruses, SARS-CoV-2 has several unique insertion sites in its spike protein (SEQ ID Nos. 50 to 55). The amino acid residues at these insertion sites in the SARS-CoV-2 S protein are identical or similar to amino acid residues in the replication protein gp120 or Gag of human immunodeficiency virus type 1 (HIV-1). In another embodiment, a series of mRNA sequences (SEQ ID Nos. 56 to 66) encoding the same amino acid residues between SARS-CoV-2 and HIV-1 were engineered, which are recognized by the human immune system.

[0037] In another implementation, the mRNA encodes the D614G variant of the ORF of the spike protein. The 2019 coronavirus disease (COVID-19) outbreak caused by SARS-CoV-2 has rapidly expanded into a global pandemic. Over time, SARS-CoV-2 isolates encoding the D614G mutation in the viral spike (S) protein have gradually become dominant where they were found, meaning this variation enhances viral transmission. S proteins with aspartic acid (D614) and glycine (G614) residues at 614 residues exhibit different functional properties. Specifically, G614 pseudotype retroviruses were found to infect ACE2-expressing cells much more efficiently than wild-type (D614) retroviruses. This greater infectivity was associated with less S1 shedding and more S protein incorporation into the pseudovirus. Similar results were obtained with virus-like particles generated from the M, N, E, and S proteins of SARS-CoV-2. In the correlation analysis, the spike protein carrying the D614G mutation showed a significant positive correlation with mortality (r = 0.43, P = 0.022). These results indicate that the spike protein carrying the D614G mutation is more stable than the wild type, consistent with epidemiological data, suggesting that the virus carrying the D614G mutation has higher transmission efficiency (SEQ ID No. 10). A vaccine containing an ORF including the D614G mutation was produced because it is more effective against the dominant strain of Sars-Cov-2.

[0038] In another embodiment, the mRNA encodes an ORF having SEQ ID NO:89, which encodes the D614G spike protein precursor S2P.

[0039] In another embodiment, the mRNA encodes an ORF having Seq ID No. 67, which incorporates other identified mutations of the spike protein. These include the HV 69-70 deletion spike protein, the Y144 deletion spike protein, the N501Y mutation spike protein, the A570D mutation spike protein, the P681H mutation spike protein, the T716I mutation spike protein, the F817P modified spike protein, the A892P modified spike protein, the A899P modified spike protein, the A942P modified spike protein, the S982A mutation spike protein, and the D1118H mutation spike protein. Vaccine compositions containing mRNA encoding one or more of these known mutations can be prepared using the methods described herein.

[0040] In other embodiments, the mRNA may encode one or more other SARS-CoV-2 viral protein-coding sequences, including envelope (E), membrane (M), or nucleocapsid (N) structural proteins. The SARS-CoV-2 genome is arranged in the sequence: 5′-replicaase (orf1 / ab)-structural protein [spike (S)-enveloping (E)-membrane (M)-nucleocapsid (N)]-3′ (Zulma et al. 2016, Nature Reviews Drug Discovery 15:327-347.), and lacks the hemagglutinin-esterase gene, which is characteristically present in the Aβ-CoV lineage. Therefore, various viral proteins other than or equal to the mRNA encoding the SARS-CoV-2 S protein can serve as targets for mRNA-based vaccination. In one embodiment, viral surface proteins are advantageously targets for mRNA vaccines. Viral functional proteins responsible for viral polymerase in the ORF1a (SEQ ID No. 5) region and replication (RdRp) in the ORF1b (SEQ ID No. 6) region can also be used to prepare vaccine compositions.

[0041] Other viral proteins can also be targets of mRNA vaccines. For example, the coronavirus membrane (M) protein plays a key role in viral assembly. One of its functions is to mediate the insertion of the spike protein into the viral envelope. The nucleocapsid N protein is a highly immunogenic phosphoprotein that is generally highly conserved. The coronavirus N protein is frequently used as a biomarker in diagnostic tests. The coronavirus (CoV) envelope (E) protein plays multiple roles during infection, including viral morphogenesis and pathogenicity. In other embodiments, mRNA encoding all or part of one or more viral M, N, and E proteins as specific antigens may be selected as components of antigenic vaccine compositions (SEQ ID No. 7, 8, 9).

[0042] The mRNA sequence can be modified to prepare vaccine compositions. For example, a 5' cap structure can be added to the mRNA sequence. In one embodiment, a 5' CleanCap is added. This structure uses an initiating capped trimer to generate a naturally occurring 5' cap structure. Advantageously, adding a multi-A tail to the sequence increases the stability of the mRNA. An AU- or GU-rich sequence can be selected in the 3' UTR, while a random sequence can be used in the 5' UTP to reduce similarity to the host gene sequence. Codon optimization of the mRNA sequence can also be used to improve translation efficiency. Specifically, codons corresponding to low-abundance tRNA species and thus resulting in lower protein expression are replaced with codons corresponding to high-abundance tRNAs that provide higher protein expression levels. (Brown, et al. Protein Expr Purif, 2008, 59:94-102; Maertens et al., 2010, Protein Sci, 9:1312-1326.; Plotkin et al., 2011, Nat Rev Genet, 12:32-42.; Kudla et al, 2009, Science, 324:255-258.).

[0043] Improve the expression efficiency of mRNA vaccines To improve mRNA expression efficiency in mammalian cells, mRNA stability can be enhanced through partial chemical modification. To further improve translation efficiency, short double-stranded RNA from aberrant RNA polymerase activity can be removed, for example, using cellulose-based FPLC, HPLC, or chromatography. To improve the potency of mRNA vaccines, sequence optimization can be used, along with modified nucleosides such as pseudouridine (φ), 5-methylcytidine (5mC), Cap-1 structures, and optimized codons, thereby improving translation efficiency. During in vitro transcription of mRNA, immature mRNA can be generated as a contaminant, which inhibits translation by stimulating innate immune activation (Kariko et al, Mol Ther. 2008 16(11):1833-40.; Andries et al, J Control Release. 2015 Nov 10;217:337-44.). FPLC and HPLC purification can be used to remove these contaminants.

[0044] In the compositions described herein, the template for in vitro transcription of mRNA contains five cis-structure elements, from 5' to 3': (i) an optimized cap structure, (ii) an optimized 5' untranslated region (UTR), (iii) a codon-optimized coding sequence, (iv) an optimized 3' UTR, and (v) a repeating adenine nucleotide (multiple A tails). (Murray et al, Mol. Cell. Biol. 2007, 27, 2791–2799. ;Louis et al, Curr. Opin. Genet. Dev. 2011, 21, 444–451. ;Ferizi et al, Lab Chip 2015, 15, 3561–3571. ;Von Niessenet et al, Mol. Ther. 2019, 27, 824–836.) These cis-structural elements are further optimized to achieve better mRNA characteristics, including half-life and expression levels. The 5'-UTR provided herein includes an initiating but non-coding polypeptide (i.e., it is non-coding). In some embodiments, the 5'-UTR of this disclosure comprises a cap structure having a 7-methylguanosine (7mG) sequence. The 3'-UTR is located directly downstream (3') of the stop codon (the codon of the mRNA transcript representing the termination signal) and does not encode a polypeptide (it is non-coding). The multi-A tail is a special region of the mRNA, located downstream of the 3'-UTR, and contains multiple consecutive adenosine monophosphates; it is required for efficient mRNA production. The mRNA in vitro transcription template is used not only for the efficiency of mRNA production but also for subsequent protein or peptide production.

[0045] In some embodiments, mRNA is produced by in vitro transcription (IVT) from a linear DNA template containing a phage promoter, an optimized UTR, and codon-optimized sequences using a mixture of RNA polymerases (T7, T3, or SP6) and different nucleosides. In other embodiments, the linear DNA template can be cloned into plasmid DNA (pDNA) as a delivery vector. Plasmid vectors can be used for the production of mRNA vaccines. Commonly used plasmids include pSFV1, pcDNA3, and pTK126, all of which are commercially available. A unique mRNA expression system is pEVL (see Grier et al. Mol Ther Nucleic Acids. 19;5:e306 (2016)).

[0046] In some embodiments, the mRNA is a modified mRNA in which all uridine residues are replaced by pseudouridine during in vitro transcription. This modification stabilizes the mRNA, prevents intracellular enzymatic degradation, and thereby improves the translation efficiency of the mRNA. The pseudouridine used can be N1-methylpseudouridine or other modifications well known in the art, such as N6-methyladenosine (m6A), inosine, pseudouridine, 5-methylcytidine (m5C), 5-hydroxymethylcytidine (hm5C), and N1-methyladenosine (m1A). This modification is optionally performed throughout the mRNA, including the ORF, 5'UTR, and 3'UTR. Those skilled in the art will recognize that other modified RNA residues can be used to stabilize the three-dimensional structure of proteins and increase protein translation.

[0047] To test the efficiency level, the experiments described in Examples 3, 4 and 5 were conducted.

[0048] Histidine-Lysine (HK) peptides as mRNA vaccine delivery systems Whether in basic research or clinical applications, effective methods for transferring nucleic acids into target cells are crucial tools. Currently, a diverse range of nucleic acid vectors is needed because the effectiveness of a specific vector depends on the characteristics of the transfected nucleic acid [Blakney et al. Biomacromolecules 2018, 19: 2870-2879. Goncalves et al. Mol Pharm 2016; 13: 3153-3163. Kauffman et al. Biomacromolecules 2018;19: 3861-3873. Peng et al. Biomacromolecules 2019; 20: 3613-3626. Scholz et al. J Control Release 2012; 161: 554-565.]. Among various vectors, non-viral delivery systems have been developed and reported to have advantages over viral delivery systems in many respects [Brito et al. Adv Genet. 2015; 89: 179-233]. For example, high molecular weight branched polyethylene glycol (PEI, 25 kDa) is an excellent vector for plasmid DNA, but not for mRNA. However, by reducing the molecular weight of PEI to 2 kDa, it becomes a more efficient vector for mRNA [Bettinger et al. Nucleic Acids Res 2001; 29: 3882-3891.].

[0049] The four-branched histidine-lysine (HK) peptide polymer H2K4b has been shown to be a good carrier for high molecular weight DNA plasmids [Leng et al. Nucleic Acids Res 2005; 33: e40.], but a poor carrier for relatively low molecular weight siRNAs [Leng et al. J Gene Med 2005; 7: 977-986.]. Two histidine-rich peptide analogs of H2K4b, namely H3K4b and H3K(+H)4b, have been shown to be effective carriers for siRNAs [Leng et al. J Gene Med 2005; 7: 977-986. Chou et al. Biomaterials 2014; 35: 846-855.], although H3K(+H)4b appears to be slightly more effective [Leng et al. Mol Ther 2012; 20: 2282-2290.]. Furthermore, the H3K4b siRNA vector induced cytokines to a significantly greater extent in vitro and in vivo than the H3K(+H)4b siRNA polymer [Leng et al. Mol Ther 2012; 20: 2282-2290.]. Suitable HK peptides are described in WO / 2001 / 047496, WO / 2003 / 090719, and WO / 2006 / 060182, the contents of which are incorporated herein by reference in their entirety. These peptides have a lysine backbone (three lysine residues) in which the ε-amino group and N-terminus of the lysine side chain are linked to various HK sequences. HK peptide vectors can be synthesized by methods well known in the art, including, for example, solid-phase synthesis.

[0050] These histidine-lysine peptide polymers (“HK polymers”) have been found to be surprisingly effective as mRNA carriers, and they can be used alone or in combination with liposomes to provide efficient delivery of mRNA into target cells. Similar to PEIs and other carriers, preliminary results indicate that HK polymers differ in their ability to carry and release nucleic acids. However, because HK polymers can be repeatedly fabricated on peptide synthesizers, their amino acid sequences can be easily altered, allowing for fine control over mRNA binding and release, as well as the stability of polymers containing HK polymers and mRNA [Chou et al. Biomaterials 2014; 35: 846-855. Midoux et al. Bioconjug Chem 1999; 10: 406-411. Henig et al. Journal of American Chemical Society 1999; 121: 5123-5126.]. When mRNA molecules are mixed with one or more HKP carriers, these components self-assemble into nanoparticles.

[0051] As described herein, advantageously, the HK polymer comprises four short peptide branches linked to a trilysine amino acid core. These peptide branches are composed of histidine and lysine amino acids in different configurations. The general structure of these histidine-lysine peptide polymers (HK polymers) is shown in Formula I, where R represents a peptide branch and K is the amino acid L-lysine.

[0052] In Formula I, K is L-lysine and each of R1, R2, R3, and R4 is independently a histidine-lysine peptide. In the HK polymer of the present invention, R 1-4 Branches can be the same or different. When an R branch is "different," the amino acid sequence of that branch differs from that of every other R branch in the polymer. Suitable R branches used in the HK polymer of the present invention shown in Formula I include, but are not limited to, the following R branches: A -R -J : R A = KHKHHKHHKHHKHHKHHKHK– (SEQ ID NO:79) R B = KHHHKHHHKHHHKHHHK– (SEQ ID NO:80) R C = KHHHKHHHKHHHHKHHHK– (SEQ ID NO:81) R D= kHHHkHHHkHHHHkHHHk– (SEQ ID NO:82) R E = HKHHHKHHHKHHHHKHHHK– (SEQ ID NO:83) R F = HHKHHHKHHHKHHHHKHHHK– (SEQ ID NO:84) R G = KHHHHKHHHHKHHHHKHHHHK– (SEQ ID NO:85) R H = KHHHKHHHKHHHKHHHHK– (SEQ ID NO:86) R I = KHHHKHHHHKHHHKHHHK– (SEQ ID NO:87) R J = KHHHKHHHHKHHHKHHHHK– (SEQ ID NO:88) Specific HK polymers that can be used in mRNA compositions include, but are not limited to, HK polymers in which each of R1, R2, R3, and R4 is identical and selected from R A - R J (Table 1). These HK polymers are named H2K4b, H3K4b, H3K(+H)4b, H3k(+H)4b, H-H3K(+H)4b, HH-H3K(+H)4b, H4K4b, H3K(1+H)4b, H3K(3+H)4b, and H3K(1,3+H)4b, respectively. In each of these 10 examples, uppercase "K" represents L-lysine, and lowercase "k" represents D-lysine. The additional histidine residues compared to H3K4b are underlined within the branched sequence. The nomenclature of HK polymers is as follows: 1) For H3K4b, the dominant repeating sequence in the branch is -HHHK-, so "H3K" is part of the name; "4b" refers to the number of branches; 2) Each branch of H3K4b and its analogues contains four -HHHK- motifs; the first -HHHK- motif ("1") is closest to the lysine core; 3) H3K(+H)4b is an analogue of H3K4b, in which an additional histidine is inserted into the second -HHHK- motif (motif 2) of H3K4b. 4) For H3K(1+H)4b and H3K(3+H)4b peptides, there are additional histidines in the first (motif 1) and third (motif 3) motifs, respectively; 5) For H3K(1,3+H)4b, there are two additional histidines in the first and third motifs of the branch.

[0053] Table 1 Well-known methods in the art, including gel lag assays, heparin replacement assays, and flow cytometry, can be used to evaluate the performance of different configurations containing HK polymers plus liposomes in successfully delivering mRNA. Suitable methods are described, for example, in Gujrati et al, | Mol.Pharmaceutics 11:2734-2744 (2014), Pärnaste et al., Mol Ther Nucleic Acids.7: 1-10 (2017).

[0054] Using SmartFlare ® The technology (Millipore Sigma) can also detect intracellular mRNA uptake. These smart signal flares are sequence-attached beads that generate increased fluorescence when an RNA sequence in a cell is detected, and this fluorescence can be analyzed using a fluorescence microscope.

[0055] Other methods include measuring protein expression from mRNA—for example, transfection efficiency can be measured using mRNA encoding luciferase using methods well known in the art. See, for example, those performed with luciferase mRNA in a recent publication (He et al, J Gene Med. 2021 Feb;23(2):e3295) to demonstrate the efficacy of delivering mRNA using HKP and liposomal formulations.

[0056] Synergistic activity of DOTAP and HK vectors in mRNA delivery The combination of H3K(+H)4b and DOTAP (cationic lipids) unexpectedly exhibited a synergistic effect in carrying mRNA into MDA-MB-231 cells (H3K(+H)4b / liposomes vs liposomes, P < 0.0001). This combination was approximately 3-fold and 8-fold more effective as mRNA carriers than the polymers and cationic lipids alone, respectively. Not all HK peptides showed synergistic activity with DOTAP lipids. For example, the combination of H3K4b and DOTAP was less effective as a carrier of luciferase mRNA than DOTAP liposomes. Besides DOTAP, other cationic lipids that can be used with HK peptides include Lipofectin (ThermoFisher), Lipofectamine (ThermoFisher), and DOSPER.

[0057] The D-isomer of H3k(+H)4b, in which the L-lysine in the branch is replaced by a D-lysine, was the most efficient polymer carrier (H3k(+H)4b vs. H3K(+H)4b, P<0.05). Compared with H3k(+H)4b alone and the liposome carrier, the D-isomer / liposome carrier of mRNA was nearly 4-fold and 10-fold more efficient, respectively. Although the D-H3k(+H)4b / liposome combination was slightly more efficient than the L-H3K(+H)4b / liposome combination, this comparison was not statistically significant.

[0058] H3K(+H)4b serves as a vector for mRNA in HK polymers. Both H3K4b and H3K(+H)4b can serve as vectors for in vitro nucleic acids [Len et al. J Gene Med 2005;7: 977-986. Chou et al., Cancer Gene Ther 2011; 18: 707-716.]. Despite these previous findings, H3K(+H)4b is significantly better as a vector for mRNA compared to other similar analogs (Table 2).

[0059] Table 2 Specifically, it exhibited higher mRNA transfection efficiency than H3K4b at various weight-to-weight (HK:mRNA) ratios. At a 4:1 ratio, H3K(+H)4b showed luciferase expression levels 10-fold higher than H3K4b in MDA-MB-231 cells without significant cytotoxicity. Furthermore, buffering capacity did not appear to be a necessary factor for their transfection differences, as the percentages of histidine (by weight) in H3K4b and H3K(+H)4b were 68.9% and 70.6%, respectively.

[0060] Gel lag assays demonstrated that the HK polymer delayed the electrophoretic migration of mRNA. The lag effect increased with increasing peptide-to-mRNA weight ratio. However, at a 2:1 ratio of H3K(+H)4b, mRNA was completely delayed, while H3K4b was not. This suggests that H3K(+H)4b can form a more stable polymer, which is advantageous for its ability to serve as a suitable carrier for mRNA delivery.

[0061] Heparin replacement assays further confirmed that the H3K(+H)4b peptide binds more tightly to mRNA. Adding different concentrations of heparin to the polymer formed by mRNA and HK revealed that, especially at low heparin concentrations, mRNA was more readily released from the H3K4b polymer than from the H3K(+H)4b polymer. These data indicate that H3K(+H)4b can bind to mRNA and form a more stable polymer than H3K4b itself.

[0062] Anthocyanin-5 labeled mRNA was used to compare the uptake of H3K4b and H3K(+H)4b polymers into MDA-MB-231 cells using flow cytometry. At different time points (1, 2, and 4 h), the H3K(+H)4b polymer was introduced into cells more efficiently than the H3K4b polymer (data not shown). Similar to these results, fluorescence microscopy showed that the H3K(+H)4b polymer was significantly more localized within acidic endosomal vesicles than the H3K4b polymer (H3K4b vs. H3K(+H)4b, P < 0.001). Interestingly, the irregularly shaped H3K4b polymers, which did not overlap with intracellular endocytic vesicles, were likely extracellular, and no H3K(+H)4b polymers were observed.

[0063] It is well known that HK polymers and cationic lipids (i.e., DOTAP) significantly and independently enhance plasmid transfection [Chen et al. Gene Ther 2000; 7: 1698-1705.]. Therefore, this study investigated whether these lipids, when combined with HK polymers, could enhance mRNA transfection. Notably, H3K(+H)4b and H3k(+H)4b vectors were significantly better mRNA carriers than DOTAP liposomes. The combination of H3K(+H)4b and DOTAP lipids showed a synergistic effect in carrying mRNA into MDA-MB-231 cells. This combination was approximately 3-fold and 8-fold more effective as mRNA carriers than polymers and liposomes alone, respectively (H3K(+H)4b / liposomes vs. liposomes or H3K(+H)4b). It is noteworthy that not all HK peptides exhibited ameliorative activity against DOTAP lipids. The combination of H3K4b and DOTAP vectors was less effective as a carrier of luciferase mRNA than DOTAP lipids. The combination of DOTAP and H3K(+H)4b vectors was found to have a synergistic effect in carrying mRNA into cells [He et al. J Gene Med. 2020 Nov 10:e3295].

[0064] mRNA vaccine administration The vaccine formulations described herein can be administered to subjects, including human subjects, via any method of administration commonly used for administering vaccines. Therefore, these compositions can be in the form of aerosols, dispersions, solutions, or suspensions and are formulated for inhalation, intramuscular, oral, sublingual, buccal, parenteral, nasal, subcutaneous, intradermal, or topical administration. The term "parenteral" as used herein includes percutaneous, subcutaneous, intravascular (e.g., intravenous), intramuscular, or intrathecal injection or infusion techniques.

[0065] As used herein, an effective dose of a vaccine refers to the dose required to produce a protective immune response in subjects receiving the vaccine. In this context, a protective immune response refers to an immune response in subjects challenged by SARS-CoV-2 that prevents or improves the disease. Methods for conducting clinical trials in multiple subjects to determine whether a vaccine produces a protective immune response are well known in the art.

[0066] The vaccine can be administered once or multiple times. A preliminary measurement of the immune response to the vaccine can be made by measuring the production of antibodies in the recipient. Methods for measuring antibody production in this manner are well known in the art. A pharmaceutically effective dose refers to the dose required to prevent, suppress, or treat (to a certain extent alleviate symptoms, preferably all symptoms) a disease state. The pharmaceutically effective dose depends on the type of disease, the composition used, the route of administration, the type of mammal being treated, the physical characteristics of the specific mammal under consideration, concurrent drug treatments, and other factors that a person skilled in the art will recognize. Generally, depending on the potency of the formulated composition, the active ingredient is administered in amounts from 0.1 mg / kg to 100 mg / kg body weight / day.

[0067] However, until recently, their application was limited by the instability and inefficiency of in vivo mRNA delivery. The methods described herein provide approaches for preparing and using mRNA vaccines with peptide delivery systems and peptide nanoparticle delivery systems.

[0068] The methods described in this article can be used for the clinical application of mRNA, including preventive and therapeutic vaccines against various diseases, especially infectious diseases.

[0069] Example 1: mRNA synthesis: mRNAs expressing CleanCap SARS-CoV 2 spike proteins (S1P and S2P) and CleanCap enhanced green fluorescent protein (EGFP) were synthesized. CleanCap SARS-CoV 2 spike protein (S6P) was prepared via TriLink BioTechnologies, Inc. (San Diego, CA). Purification was achieved by selective binding of dsRNA to cellulose in ethanol-containing buffer A chromatography buffer containing 10 mM HEPES (pH 7.2), 0.1 mM EDTA, 125 mM NaCl, and 16% ethanol. The cellulose cellulose and mRNA product were added together to a microcentrifuge column and centrifuged. After this process, nearly 90% of the dsRNA was removed (Baiersdorfer et al., 2019). Contaminants could also be eliminated by FPLC and HPLC (Kariko et al., 2011).

[0070] Example 2: Preparation of peptides (HK polymers) HK peptide polymers were synthesized on the RaininVoyager synthesizer (Tucson, AZ) using the Biopolymer Core Facility at the University of Maryland.

[0071] Example 3: In vitro transfection of mRNA To validate appropriate protein expression of the mRNAs (EGFP mRNA and SARS-CoV 2 SP), the mRNAs were transiently transfected into human embryonic kidney 293T cells (293T cells). In short, 4.8 x 10⁻⁶ mRNAs were transfected into these cells. 5 Cells were seeded into 6-well plates containing 2 mL of DMEM (10% fetal bovine serum and 1% penicillin-streptomycin (ThermoFisher Scientific)). After 24 hours, when the cells were 70-90% confluent, mRNA was transfected into 293T cells using the Lipofectamine MessengerMAX transfection reagent (ThermoFisher Scientific) according to the provided experimental protocol. 293T cells were cultured for two days, and in vitro protein expression was measured.

[0072] The ability of two HK-based polymers carrying EGFP mRNA and SARS-CoV 2 SP mRNA, bound to cationic liposomes DOTAP (1,2-dioleoyl-3-trimethylammonium-propane, Roche), MC3 (DLin-MC3-DMA, ChemScene), and PLA (polylactic acid) to enter human embryonic kidney 293 cells (HEK 293 cells) was examined. Briefly, 4.8 x 10⁻⁶ cells were transported to the target cells. 5 Cells were seeded into 6-well plates containing 2 mL of DMEM (10% fetal bovine serum and penicillin-streptomycin (ThermoFisher Scientific)). After 24 hours, when the cells reached 70-90% confluence, two HK-based polymer formulations containing EGFP and SP mRNA were added to each well. 293T cells were cultured for two days, and in vitro protein expression measurements were performed.

[0073] Example 4: In vitro protein expression measurement Immunofluorescence analysis: Two days after transfection, protein expression was measured by immunofluorescence imaging using a Cytation 5 Cell Imaging Multi-Mode Reader (Biotek, Winooski, VT).

[0074] Preparation of cell lysis buffer: Two days after transfection, the culture medium was aspirated, and the cells were washed with ice-cold PBS on ice. Ice-cold lysis buffer (RIPA, ThermoFisher Scientific) containing a protease inhibitor (ThermoFisher Scientific) was added, and the cells were incubated at 4˚C for 30 minutes. Cells were harvested with a cell scraper and lysed by sonication. The cells were centrifuged at 10,000g for 20 minutes at 4°C to precipitate cell debris, and the supernatant was transferred to a new microcentrifuge tube. The protein concentration of the lysis buffer was determined using Bradford or BCA protein assays for Western blotting.

[0075] Western blotting: Simply put, 20-50 µg of protein is loaded into each well of the gel along with 4X SDS sample buffer (ThermoFisher Scientific), 10X reduction buffer (ThermoFisher Scientific), and an additional ddH2O (ThermoFisher Scientific), for a total loading of 25 µl / well. The mixture is denatured at 95 °C for 5 min, cooled to room temperature, centrifuged, and then loaded into a NuPAGE™ 4-12% Bis-Tris gel (ThermoFisher Scientific). After electrophoresis separation, the gel is removed from the cartridge and transferred using the iBlot™ 2 dry blot system (ThermoFisher Scientific). The transfer membrane was blocked with 5% nonfat milk powder in TBST for 1 hour at RT, incubated overnight with primary antibody at 4°C, washed three times with TBST (0.05% Tween 20 in TBS) buffer, and then incubated with secondary antibody (HRP-linked mouse IgG (H+L) secondary antibody, ThermoFisher Scientific, A24512) for 1 hour at RT. The transfer membrane was developed using PierceECL Western Blotting Substrate (ThermoFisher Scientific) and imaged using a chemiluminescence imaging system.

[0076] Example 5: Preparation of in vivo formulation 1. PNI-Genvoy LNP formulation: Lipid nanoparticles were formulated using the GenVoy platform with PNI NanoAssemblr (Precision NanoSystems, Vancouver, British Columbia, Canada) as a positive control for in vitro and in vivo experiments.

[0077] 2. HKP(+H) Preparation: Prepare a stock solution of HKP(+H) (10 mg / mL) in nuclease-free water. Dilute the concentrated stock solution to 4 mg / mL in water. Prepare a working mRNA solution (1 mg / mL) in 1 mM citrate buffer (pH 6.0). Form an mRNA / HKP(+H) polymer by mixing equal volumes of 4 mg / mL HKP(+H) and 1 mg / mL mRNA. The mass ratio of HKP(+H) to mRNA is 4:1. Incubate the mRNA / HKP(+H) polymer at room temperature for 30 minutes before use. Determine the size of each peptide polymer using a Zetasizer (Malvern Panalytical) before transfection or injection.

[0078] 3. HKP(+H) / DOTAP formulation (with DOTAP added later): Prepare a stock solution of HKP(+H) (10 mg / mL) in nuclease-free water. Dilute the concentrated stock solution to 4 mg / mL in water. DOTAP (Sigma-Aldrich) is 1 mg / mL in an aqueous buffer. Prepare a working solution of mRNA (1 mg / mL) in 1 mM citrate buffer (pH 6.0). First, form an mRNA / HKP(+H) polymer by mixing equal volumes of 4 mg / mL HKP(+H) and 1 mg / mL mRNA. Incubate the mRNA / HKP(+H) polymer at room temperature for 30 minutes. Next, add equal volumes of DOTAP and HKP(+H) solution to the mRNA / HKP(+H) polymer. The mass ratio of HKP(+H) / DOTAP to mRNA is 4:1:1. Incubate the mRNA / HKP(+H) / DOTAP nanoparticles at room temperature for 30 minutes before use.

[0079] 4. HKP(+H) / MC3 or HKP(+H) / DOTAP formulation (premixed MC3 or DOTAP): Prepare a stock solution of HKP(+H) (10 mg / mL) in nuclease-free water. Dilute the concentrated stock solution to 4 mg / mL in water. DOTAP or MC3 is 1 mg / mL in buffered aqueous solution. Prepare a working mRNA solution (1 mg / mL) in 1 mM citrate buffer (pH 6.0). Premix equal volumes of HKP(+H) and MC3 at a 4:1 mass ratio, and add an equal volume of mRNA and HKP(+H) solution to the premixed HKP(+H) / MC3. Mix the premixed 4 mg / mL HKP(+H) / 1 mg / mL MC3 and 1 mg / mL mRNA to form mRNA / HKP(+H) / MC3 nanoparticles. The mass ratio of HKP(+H) / MC3 to mRNA is 4:1:1. Incubate the mRNA / HKP(+H) / MC3 nanoparticles at room temperature for 30 minutes before use.

[0080] 5. HKP(+H) / PLA NP formulation: Prepare a stock solution of HKP(+H) (10 mg / mL) in nuclease-free water. Dilute the concentrated stock solution to 4 mg / mL in water. Prepare poly-L-lactic acid (PLA) nanoparticles (5 mg / mL) in water. Prepare a working solution of mRNA (1 mg / mL) in 1 mM citrate buffer (pH 6.0). Mix equal volumes of HKP(+H) and mRNA at a mass ratio of 4:1. Incubate the mRNA / HKP(+H) polymer at room temperature for 30 minutes, then add an equal volume of PLA nanoparticles and HKP(+H) solution to the mRNA / HKP(+H) nanoparticles, allowing the mRNA / HKP(+H) polymer to be adsorbed onto the surface of the PLA nanoparticles. The mass ratio of HKP(+H) / PLA to mRNA is 4:5:1. Incubate the mRNA / HKP(+H) / PLA nanoparticles at room temperature for 30 minutes before use.

[0081] Example 6: In vivo animal model and injection In vivo studies were conducted at Antibody and Immunoassay Consultants (AIC) Inc (Gaithersburg, MD). Briefly, eight 6-8 week old female BALB / c mice were randomly assigned to groups of four, and each group received an intramuscular injection of 30 µg of different formulations of EGFP into the right abdomen. CleanCap SARS-CoV 2 spike protein (S6P) polymers were prepared using the same formulations for in vivo analysis and antibody titer and binding assays.

[0082] A second injection was given on day 28 to enhance immunity, and serum was collected on day 35 and analyzed using an immunoassay (ELISA) to measure antibody titers.

[0083] result Immunofluorescence analysis One week after preparation, the formulation was delivered to HEK 293T cells. Figure 2 All HKP(H) groups demonstrated the possibility of carrying mRNA into cells in vitro. Within the HKP(H) groups, the HKP(H)-only group showed higher EGFP expression than other groups and the PNI Genvoy (positive control).

[0084] Western blot analysis Next, Western blot analysis was performed, and the results were ( Figure 3 The results showed that the spike protein expressed on the cell surface by the S1P D614G mRNA delivery group detached after localization to the cell membrane. Only the S2 domain of the spike protein remained on the cell surface. The band size of the protein blot (70–80 kDa) supported this hypothesis. However, the spike protein of S2P D614G showed the full size of the spike protein (180 kDa). sequence list <110> Da Mian Biotechnology Co., Ltd. <120> Composition of mRNA vaccine against novel coronavirus infection <130> 3949.0001I <140> PCT / US2021 / 017149 <141> 2021-02-08 <150> 63 / 130,581 <151> 2020-12-24 <150> 63 / 058,463 <151> 2020-07-29 <150> 62 / 971,834 <151> 2020-02-07 <160> 108 <170> PatentIn version 3.5 <210> 1 <211> 1518 <212> DNA <213> Severe acute respiratory syndrome coronavirus 2 <400> 1 atgttcttgt taacaactaa acgaacaatg tttgtttttc ttgttttatt gccactagtc 60 tctagtcagt gtgttaatct tacaaccaga actcaattac cccctgcata cactaattct 120 ttcacacgtg gtgtttatta ccctgacaaa gttttcagat cctcagtttt acattcaact 180 caggacttgt tcttaccttt cttttccaat gttacttggt tccatgctat acatgtctct 240 gggaccaatg gtactaagag gtttgataac cctgtcctac catttaatga tggtgtttat 300 tttgcttcca ctgagaagtc taacataata agaggctgga tttttggtac tactttagat 360 tcgaagaccc agtccctact tattgttaat aacgctacta atgttgttat taaagtctgt 420 gaatttcaat tttgtaatga tccatttttg ggtgtttatt accacaaaaa caacaaaagt 480 tggatggaaa gtgagttcag agtttattct agtgcgaata attgcacttt tgaatatgtc 540 tctcagcctt ttcttatgga ccttgaagga aaacagggta atttcaaaaa tcttagggaa 600 tttgtgttta agaatattga tggttatttt aaaatatatt ctaagcacac gcctattaat 660 ttagtgcgtg atctccctca gggtttttcg gctttagaac cattggtaga tttgccaata 720 ggtattaaca tcactaggtt tcaaacttta cttgctttac atagaagtta tttgactcct 780 ggtgattctt cttcaggttg gacagctggt gctgcagctt attatgtggg ttatcttcaa 840 cctaggactt ttctattaaa atataatgaa aatggaacca ttacagatgc tgtagactgt 900 gcacttgacc ctctctcaga aacaaagtgt acgttgaaat ccttcactgt agaaaaagga 960 atctatcaaa cttctaactt tagagtccaa ccaacagaat ctattgttag atttcctaat 1020 attacaaact tgtgcccttt tggtgaagtt tttaacgcca ccagatttgc atctgtttat 1080 gcttggaaca ggaagagaat cagcaactgt gttgctgatt attctgtcct atataattcc 1140 gcatcatttt ccacttttaa gtgttatgga gtgtctccta ctaaattaaa tgatctctgc 1200 tttactaatg tctatgcaga ttcatttgta attagaggtg atgaagtcag acaaatcgct 1260 ccagggcaaa ctggaaagat tgctgattat aattataaat taccagatga tttacaggc 1320 tgcgttatag cttggaattc taacaatctt gattctaagg ttggtggtaa ttataattac 1380 ctgtatagat tgtttaggaa gtctaatctc aaaccttttg agagagatat ttcaactgaa 1440 atctatcagg ccggtagcac accttgtaat ggtgttgaag gttttaattg ttactttcct 1500 ttacaatcat atggtttc 1518 <210> 2 <211> 700 <212> DNA <213> Severe acute respiratory syndrome coronavirus 2 <400> 2 caacccacta atggtgttgg ttaccaacca tacagagtag tagtactttc ttttgaactt 60 ctacatgcac cagcaactgt ttgtggacct aaaaagtcta ctaatttggt taaaaacaaa 120 tgtgtcaatt tcaacttcaa tggtttaaca ggcacaggtg ttcttactga gtctaacaaa 180 aagttctgc ctttccaaca atttggcaga gacattgctg acactactga tgctgtccgt 240 gatccacaga cacttgagat tcttgacatt acaccatgtt cttttggtgg tgtcagtgtt 300 ataacaccag gaacaaatac ttctaaccag gttgctgttc tttatcagga tgttaactgc 360 acagaagtcc ctgttgctat tcatgcagat caacttactc ctacttggcg tgtttattct 420 acaggttcta atgttttca aacacgtgca ggctgtttaa taggggctga acatgtcaac 480 aactcatatg agtgtgacat acccattggt gcaggtatat gcgctagtta tcagactcag 540 actaattctc ctcggcgggc acgtagtgta gctagtcaat ccatcattgc ctacactatg 600 tcacttggtg cagaaaattc agttgcttac tctaataact ctattgccat acccacaaat 660 tttactatta gtgttaccac agaaattcta ccagtgtcta 700 <210> 3 <211> 800 <212> DNA <213> Severe acute respiratory syndrome coronavirus 2 <400> 3 tgaccaagac atcagtagat tgtacaatgt acatttgtgg tgattcaact gaatgcagca 60 atcttttgtt gcaatatggc agtttttgta cacaattaaa ccgtgcttta actggaatag 120 ctgttgaaca agacaaaaac acccaagaag tttttgcaca agtcaaacaa atttacaaaa 180 caccaccaat taaagatttt ggtggtttta atttttcaca aatattacca gatccatcaa 240 aaccaagcaa gaggtcattt attgaagatc tacttttcaa caaagtgaca cttgcagatg 300 ctggcttcat caaacaatat ggtgattgcc ttggtgatat tgctgctaga gacctcattt 360 gtgcacaaaa gtttaacggc cttactgttt tgccaccttt gctcacagat gaaatgattg 420 ctcaatacac ttctgcactg ttagcgggta caatcacttc tggttggacc ttggtgcag 480 gtgctgcatt acaaatacca tttgctatgc aaatggctta taggtttaat ggtattggag 540 ttacacagaa tgttctctat gagaaccaaa aattgattgc caaccaattt aatagtgcta 600 ttggcaaaat tcaagactca cttcttcca cagcaagtgc acttggaaaa cttcaagatg 660 tggtcaacca aaatgcacaa gctttaaaca cgcttgttaa acaacttagc tccaatttg 720 gtgcaatttc aagtgtttta aatgatatcc tttcacgtct tgacaaagtt gaggctgaag 780 tgcaaattga taggttgatc 800 <210> 4 <211> 831 <212> DNA <213> Severe acute respiratory syndrome coronavirus 2 <400> 4 acaggcagac ttcaaagtttt gcagacatat gtgactcaac aattaattag agctgcagaa 60 atcagagctt ctgctaatct tgctgctact aaaatgtcag agtgtgtact tggacaatca 120 aaaagagttg atttttgtgg aaagggctat catcttatgt ccttccctca gtcagcacct 180 catggtgtag tcttcttgca tgtgacttat gtccctgcac aagaaaagaa cttcacaact 240 gctcctgcca tttgtcatga tggaaaagca cactttcctc gtgaaggtgt ctttgtttca 300 aatggcacac actggtttgt aacacaaagg aatttttatg aaccacaaat cattactaca 360 gacaacacat ttgtgtctgg taactgtgat gttgtaatag gaattgtcaa caacacagtt 420 tatgatcctt tgcaacctga attagactca ttcaaggagg agttagataa atattttaag 480 aatcatacat caccagatgt tgatttaggt gacatctctg gcattaatgc ttcagttgta 540 aacattcaaa aagaaattga ccgcctcaat gaggttgcca agaatttaaa tgaatctctc 600 atcgatctcc aagaacttgg aaagtatgag cagtatataa aatggccatg gtacatttgg 660 ctaggtttta tagctggctt gattgccata gtaatggtga caattatgct ttgctgtatg 720 accagttgct gtagttgtct caagggctgt tgttcttgtg gatcctgctg caaatttgat 780 gaagacgact ctgagccagt gctcaaagga gtcaaattac attacacata a 831 <210> 5 <211> 888 <212> DNA <213> Severe acute respiratory syndrome coronavirus 2 <400> 5 gccacgtata aaccaaatac ctggtgtata cgttgtcttt ggagcacaaa accagttgaa 60 acatcaaatt cgtttgatgt actgaagtca gaggacgcgc agggaatgga taatcttgcc 120 tgcgaagatc taaaccagt ctctgaagaa gtagtggaaa atcctaccat acagaaagac 180 gttcttgagt gtaatgtgaa aactaccgaa gttgtaggag acattatact taaaccagca 240 aataatagtt taaaattac agaagaggtt ggccacacag atctaatggc tgcttatgta 300 gacaattcta gtcttactat taagaaacct aatgaattat ctagagtatt aggtttgaaa 360 acccttgcta ctcatggttt agctgctgtt aatagtgtcc cttgggatac tatagctaat 420 tatgctaagc cttttcttaa caaagttgtt agtacaacta ctaacatagt tacacggtgt 480 ttaaaccgtg tttgtactaa ttatatgcct tatttcttta ctttattgct acaattgtgt 540 acttttacta gaagtacaaa ttctagaatt aaagcatcta tgccgactac tatagcaaag 600 aatactgtta agagtgtcgg taaatttgt ctagaggctt catttaatta tttgaagtca 660 cctaattttt ctaaactgat aaattata atttggtttt tactattaag tgtttgccta 720 ggttctttaa tctactcaac cgctgcttta ggtgttttaa tgtctaattt aggcatgcct 780 840. tcttactgta ctggttacag agaaggctat ttgaactcta ctaatgtcac tattgcaacc tactgtactg gttctatacc ttgtagtgtt tgtcttagtg gtttag 888 <210> 6 <211> 901 <212> DNA <213> Severe acute respiratory syndrome coronavirus <400> 6 60. tagaaaccc tcaccttatg ggttggggatt atcctaaatg tgatagcc atgcctaaca 120. tgcttagaat tatggcctca cttgttcttg ctcgcaaaca tacaacgtgt tgtagcttgt cacaccgttt ctatagatta gctaatgagt gtgctcaagt attgagtgaa atggtcatgt gtggcggttc actatatgtt aaaccaggtg gaacctcatc aggagatgcc acaactgctt 240 atgctaatag tgtttttaac atttgtcaag ctgtcacggc caatgttaat gcacttttat ctactgatgg taacaaaatt gccgataagt atgtccgcaa tttacaacac agactttatg agtgtctcta tagaaataga gatgttgaca cagactttgt gaatgagttt tacgcatatt tgcgtaaaca tttctcaatg atgatactct ctgacgatgc tgttgtgtgt ttcaatagca 480 cttatgcatc tcaaggtcta gtggctagca taaagaactt taagtcagtt ctttattatc 540 aaaacaatgt tttatgtct gaagcaaaat gttggactga gactgacctt actaaaggac 600 ctcatgaatt ttgctctcaa catacaatgc tagttaaaca gggtgatgat tatgtgtacc 660 ttccttaccc agatccatca agaatcctag gggccggctg ttttgtagat gatatcgtaa 720 aaacagatgg tacacttatg attgaacggt tcgtgtcttt agctatagat gcttacccac 780 ttactaaaca tcctaatcag gagtatgctg atgtctttca tttgtactta caatacataa 840 gaaagctaca tgatgagtta acaggacaca tgttagacat gtattctgtt atgcttacta 900 to 901 <210> 7 <211> 669 <212> DNA <213> Severe acute respiratory syndrome coronavirus 2 <400> 7 atggcagatt ccaacggtac tattaccgtt gaagagctta aaaagctcct tgaacaatgg 60 aacctagtaa taggtttcct attccttaca tggatttgtc ttctacaatt tgcctatgcc 120 aacaggaata ggtttttgta tataattaag ttaattttcc tctggctgtt atggccagta 180 actttagctt gttttgtgct tgctgctgtt tacagaataa attggatcac cggtggaatt 240 gctatcgcaa tggcttgtct tgtaggcttg atgtggctca gctacttcat tgcttctttc 300 agactgtttg cgcgtacgcg ttccatgtgg tcattcaatc cagaaactaa cattcttctc 360 aacgtgccac tccatggcac tattctgacc agaccgcttc tagaaagtga actcgtaatc 420 ggagctgtga tccttcgtgg acatcttcgt attgctggac accatctagg acgctgtgac 480 atcaaggacc tgcctaaaga aatcactgtt gctacatcac gaacgctttc ttattacaaa 540 ttgggagctt cgcagcgtgt agcaggtgac tcaggttttg ctgcatacag tcgctacagg 600 attggcaact ataaattaaa cacagaccat tccagtagca gtgacaatat tgctttgctt 660 gtacagtaa 669 <210> 8 <211> 1260 <212> DNA <213> Severe acute respiratory syndrome coronavirus 2 <400> 8 atgtctgata atggacccca aaatcagcga aatgcacccc gcattacgtt tggtggaccc 60 tcagattcaa ctggcagtaa ccagaatgga gaacgcagtg gggcgcgatc aaaacaacgt 120 cggccccaag gtttacccaa taatactgcg tcttggttca ccgctctcac tcaacatggc 180 aaagaagacc ttaaattccc tcgaggacaa ggcgttccaa ttaacaccaa tagcagtcca 240 gatgaccaaa ttggctacta ccgaagagct accagacgaa ttcgtggtgg tgacggtaaa 300 atgaaagatc tcagtccaag atggtatttc tactacctag gaactgggcc agaagctgga 360 cttccctatg gtgctaacaa agacggcatc atatgggttg caactgaggg agccttgaat 420 acaccaaaag atcacattgg cacccgcaat cctgctaaca atgctgcaat cgtgctacaa 480 cttcctcaag gaacaacatt gccaaaaggc ttctacgcag aagggagcag aggcggcagt 540 caagcctctt ctcgttcctc atcacgtagt cgcaacagtt caagaaattc aactccaggc 600 agcagtaggg gaacttctcc tgctagaatg gctggcaatg gcggtgatgc tgctcttgct 660 ttgctgctgc ttgacagatt gaaccagctt gagagcaaaa tgtctggtaa aggccaacaa 720 caacaaggcc aaactgtcac taagaaatct gctgctgagg cttctaagaa gcctcggcaa 780 aaacgtactg ccactaaagc atacaatgta acacaagctt tcggcagacg tggtccagaa 840 caacccaag gaatttttgg ggaccaggaa ctaatcagac aaggaactga ttacaacat 900 tggccgcaaa ttgcacaatt tgccccagc gcttcagcgt tctcggaat gtcgcgcatt 960 ggcatggaag tcacaccttc gggaacgtgg ttgacctaca caggtgccat caaattggat 1020 ghaaagatc caatttca nationalcaagtc atttgctga ataagcatat tgacgcatac 1080 aaaacattcc caccaacaga gcctaaaag ghaaaaaga agaaggctga tgaaactca 1140 gccttaccgc agagacagaa gaacagcaa actgtgactc ttctcctgc tgcagatttg 1200 gatgatttct gcaacaatcc atgagcagtg ctgactcaac tcaggccta 1260 <210> 9 <211> 228 <212> DNA <213> Severe acute respiratory syndrome coronavirus <400> 9 atgtactcat tcgtttcgga agagacaggt acgttaatag ttaatagcgt acttcttttt 60 cttgctttcg tggtattctt gctagttaca ctagccatcc ttactgcgct tcgattgtgt 120 gcgtactgct gcaatattgt taacgtgagt cttgtaaaac cttcttttta cgtttactct 180 cgtgttaaaa atctgaattc ttctagagtt cctgatcttc tggtctaa 228 <210> 10 <211> 3822 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 10 atgtttgttt ttcttgtttt attgccacta gtctctagtc agtgtgttaa tcttacaacc 60 agaactcaat taccccctgc atacactaat tctttcacac gtggtgttta ttaccctgac 120 aaagttttca gatcctcagt tttacattca actcaggact tgttcttacc tttcttttcc 180 aatgttactt ggttccatgc tatacatgtc tctgggacca atggtactaa gaggtttgat 240 aaccctgtcc taccatttaa tgatggtgtt tattttgctt ccactgagaa gtctaacata 300 ataagaggct ggatttttgg tactacttta gattcgaaga cccagtccct acttattgtt 360 aataacgcta ctaatgttgt tattaaagtc tgtgaatttc aattttgtaa tgatccattt 420 ttgggtgtttt attaccacaaaacaaaa agttggatgg aaagtgagtt cagagtttat 480 tctagtgcga atattgcac ttttgaatat gtctctcagc cttttcttat ggaccttgaa 540 ggaaaacagg gtaatttca aaatcttagg gatttgtgt ttaagaat tgatggttat 600 tttaaaatat attctaagca cacgcctatt aatttagtgc gtgatctccc tcagggtttt 660 tcggctttag aaccattggt agatttgcca ataggtatta acatcactag gtttcaact 720 ttacttgctt tacatagaag tatttgact cctggtgat cttctcagg ttggacagct 780 ggtgctgcag cttattatgt gggttatctt caactagga ctttctatt aaaatataat 840 gaaaatggaa ccattacaga tgctgtagac tgtgcacttg accctctc agaaacaaag 900 tgtacgttga atccttcac tgtagaaaaa ggaatctatc aaacttctaa ctttagagtc 960 caccacag aatctattgt tagattcct atattacaa acttgtgccc ttttggtgaa 1020 gtttttaacg ccaccagatt tgcatctgtt tatgcttgga acaggaagg atcagcaac 1080 tgtgttgctg attattctgt cctatataat tccgcatcat tttccactt taagtgttat 1140 ggagtgtctc ctactaaatt aaatgatctc tgctttacta atgtctatgc agattcattt 1200 gtaattagag gtgatgaagt cagacaaatc gctccagggc aaactggaaa gattgctgat 1260 tataattata aattaccaga tgattttaca ggctgcgtta tagcttggaa ttctaacaat 1320 cttgattcta aggttggtgg taattataat tacctgtata gattgtttag gaagtctaat 1380 ctcaaacctt ttgagagaga tatttcaact gaaatctatc aggccggtag cacaccttgt 1440 aatggtgttg aaggttttaa ttgttacttt cctttacaat catatggttt ccaacccact 1500 aatggtgttg gttaccaacc atacagagta gtagtacttt cttttgaact tctacatgca 1560 ccagcaactg tttgtggacc taaaaagtct actaatttgg ttaaaaacaa atgtgtcaat 1620 ttcaacttca atggtttaac aggcacaggt gttcttactg agtctaacaa aaagtttctg 1680 cctttccaac aatttggcag agacattgct gacactactg atgctgtccg tgatccacag 1740 acacttgaga ttcttgacat tacaccatgt tctttttggtg gtgtcagtgt tataacacca 1800 ggaacaaata cttctaacca ggttgctgtt ctttatcagg gtgttaactg cacagaagtc 1860 cctgttgcta ttcatgcaga tcaacttact cctacttggc gtgtttattc tacaggttct 1920 aatgtttttc aaacacgtgc aggctgttta ataggggctg aacatgtcaa caactcatat 1980 gagtgtgaca tacccattgg tgcaggtata tgcgctagtt atcagactca gactaattct 2040 cctcggcggg cacgtagtgt agctagtcaa tccatcattg cctacactat gtcacttggt 2100 gcagaaaatt cagttgctta ctctaataac tctattgcca tacccacaaa ttttactatt 2160 agtgttacca cagaaattct accagtgtct atgaccaaga catcagtaga ttgtacaatg 2220 tacatttgtg gtgattcaac tgaatgcagc aatcttttgt tgcaatatgg cagtttttgt 2280 acacaattaa accgtgcttt aactggaata gctgttgaac aagacaaaaa cacccaagaa 2340 gtttttgcac aagtcaaaca aatttacaaa acaccaccaa ttaaagattt tggtggtttt 2400 aatttttcac aaatattacc agatccatca aaaccaagca agaggtcatt tattgaagat 2460 ctacttttca acaaagtgac acttgcagat gctggcttca tcaaacaata tggtgattgc 2520 cttggtgata ttgctgctag agacctcatt tgtgcacaaa agtttaacgg ccttactgtt 2580 ttgccacctt tgctcacaga tgaaatgatt gctcaataca cttctgcact gttagcgggt 2640 acaatcactt ctggttggac ctttggtgca ggtgctgcat tacaaatacc atttgctatg 2700 caaatggctt ataggtttaa tggtattgga gttacacaga atgttctcta tgagaaccaa 2760 aaattgattg ccaaccaatt taatagtgct attggcaaaa ttcaagactc actttcttcc 2820 acagcaagtg cacttggaaa acttcaagat gtggtcaacc aaaatgcaca agctttaaac 2880 acgcttgtta aacaacttag ctccaatttt ggtgcaattt caagtgtttt aaatgatatc 2940 ctttcacgtc ttgacaaagt tgaggctgaa gtgcaaattg ataggttgat cacaggcaga 3000 cttcaaagtt tgcagacata tgtgactcaa caattaatta gagctgcaga aatcagagct 3060 tctgctaatc ttgctgctac taaaatgtca gagtgtgtac ttggacaatc aaaaagagtt 3120 gatttttgtg gaaagggcta tcatcttatg tccttccctc agtcagcacc tcatggtgta 3180 gtcttcttgc atgtgactta tgtccctgca caagaaaaga acttcacaac tgctcctgcc 3240 atttgtcatg atggaaaagc acactttcct cgtgaaggtg tctttgtttc aaatggcaca 3300 cactggtttg taacacaaag gaatttttat gaaccacaaa tcattactac agacaacaca 3360 tttgtgtctg gtaactgtga tgttgtaata ggaattgtca acaacacagt ttatgatcct 3420 ttgcaacctg aattagactc attcaaggag gagttagata aatattttaa gaatcataca 3480 tcaccagatg ttgatttagg tgacatctct ggcattaatg cttcagttgt aaacattcaa 3540 aaagaaattg accgcctcaa tgaggttgcc aagaatttaa atgaatctct catcgatctc 3600 caagaacttg gaaagtatga gcagtatata aaatggccat ggtacatttg gctaggtttt 3660 atagctggct tgattgccat agtaatggtg acaattatgc tttgctgtat gaccagttgc 3720 tgtagttgtc tcaagggctg ttgttcttgt ggatcctgct gcaaatttga tgaagacgac 3780 tctgagccag tgctcaaagg agtcaaatta cattacacat aa 3822 <210> 11 <211> 1273 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 11 Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val 1 5 10 15 Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe 20 25 30 Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu 35 40 45 His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp 50 55 60 Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp 65 70 75 80 Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu 85 90 95 Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser 100 105 110 Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile 115 120 125 Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr 130 135 140 Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr 145 150 155 160 Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu 165 170 175 Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe 180 185 190 Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr 195 200 205 Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu 210 215 220 Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr 225 230 235 240 Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser 245 250 255 Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro 260 265 270 Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala 275 280 285 Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys 290 295 300 Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val 305 310 315 320 Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys 325 330 335 Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala 340 345 350 Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu 355 360 365 Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro 370 375 380 Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe 385 390 395 400 Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly 405 410 415 Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys 420 425 430 Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn 435 440 445 Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe 450 455 460 Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys 465 470 475 480 Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly 485 490 495 Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val 500 505 510 Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys 515 520 525 Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn 530 535 540 Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu 545 550 555 560 Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val 565 570 575 Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe 580 585 590 Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val 595 600 605 Ala Val Leu Tyr Gln Gly Val Asn Cys Thr Glu Val Pro Val Ala Ile 610 615 620 His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser 625 630 635 640 Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val 645 650 655 Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala 660 665 670 Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val Ala 675 680 685 Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser 690 695 700 Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile 705 710 715 720 Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val 725 730 735 Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu 740 745 750 Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr 755 760 765 Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln 770 775 780 Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe 785 790 795 800 Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser 805 810 815 Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly 820 825 830 Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp 835 840 845 Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu 850 855 860 Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly 865 870 875 880 Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile 885 890 895 Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr 900 905 910 Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn 915 920 925 Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala 930 935 940 Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn 945 950 955 960 Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val 965 970 975 Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln 980 985 990 Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val 995 1000 1005 Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn 1010 1015 1020 Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys 1025 1030 1035 Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro 1040 1045 1050 Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val 1055 1060 1065 Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His 1070 1075 1080 Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn 1085 1090 1095 Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln 1100 1105 1110 Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val 1115 1120 1125 Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro 1130 1135 1140 Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn 1145 1150 1155 His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn 1160 1165 1170 Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu 1175 1180 1185 Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu 1190 1195 1200 Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu 1205 1210 1215 Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met 1220 1225 1230 Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys 1235 1240 1245 Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro 1250 1255 1260 Val Leu Lys Gly Val Lys Leu His Tyr Thr 1265 1270 <210> 12 <211> 1132 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 12 attaatacga ctcactataa ggaggccacc atgttcgtgt ttctggtgct gctgcctctg 60 gtgtccagcc agcgggtgca gcccaccgaa tccatcgtgc ggttccccaa tatcaccaat 120 ctgtgcccct tcggcgaggt gttcaatgcc accagattcg cctctgtgta cgcctggaac 180 cggaagcgga tcagcaattg cgtggccgac tactccgtgc tgtacaactc cgccagcttc 240 agcaccttca agtgctacgg cgtgtcccct accaagctga acgacctgtg cttcacaaac 300 gtgtacgccg acagcttcgt gatccgggga gatgaagtgc ggcagattgc ccctggacag 360 acaggcaaga tcgccgacta caactacaag ctgcccgacg acttcaccgg ctgtgtgatt 420 gcctggaaca gcaacaacct ggactccaaa gtcggcggca actacaatta cctgtaccgg 480 ctgttccgga agtccaatct gaagcccttc gagcgggaca tctccaccga gatctatcag 540 gccggcagca ccccttgtaa cggcgtggaa ggcttcaact gctacttccc actgcagtcc 600 tacggctttc agcccacaaa tggcgtgggc tatcagccct acagagtggt ggtgctgagc 660 ttcgaactgc tgcatgcccc tgccacagtg tgcggcccta agaaaagcac caatctcgtg 720 aagaacaaat gcgtgaactt cggatccgga ggtggataca tcccggaggc ccctagggac 780 ggtcaagctt acgtgagaaa ggacggcgaa tgggttctgc tgtcgacctt cttgggataa 840 gcaagctgct tacatagaac tcgcggcgat tggcatgccg ccttaaaatt tttattttat 900 ttttcttt cttttccgaa tcggattttg ttttaatat ttcaaaaaaa aaaaaaaaaa 960 aaaaaaaacg cgaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaaa 1020 attaataaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaa 1080 ataaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaa aaaaaaaaaa aa 1132 <210> 13 <211> 269 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 13 Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Arg Val 1 5 10 15 Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys 20 25 30 Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala 35 40 45 Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu 50 55 60 Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro 65 70 75 80 Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe 85 90 95 Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly 100 105 110 Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys 115 120 125 Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn 130 135 140 Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe 145 150 155 160 Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys 165 170 175 Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly 180 185 190 Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val 195 200 205 Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys 210 215 220 Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Gly Ser Gly 225 230 235 240 Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg 245 250 255 Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu Gly 260 265 <210> 14 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 14 Arg Asp Ile Ala Asp Thr Thr Asp Ala Val Arg Asp Pro Gln 1 5 10 <210> 15 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 15 Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp 1 5 10 <210> 16 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 16 Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala Arg Ser Val 1 5 10 <210> 17 <211> 13 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 17 Leu Thr Pro Gly Asp Ser Ser Ser Gly Trp Thr Ala Gly 1 5 10 <210> 18 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 18 Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe 1 5 10 <210> 19 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 19 Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser 1 5 10 <210> 20 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 20 Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln 1 5 10 <210> 21 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 21 Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly 1 5 10 <210> 22 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 22 Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val 1 5 10 <210> 23 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 23 Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp Asn 1 5 10 <210> 24 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 24 Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser 1 5 10 <210> 25 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 25 Phe Asp Glu Asp Asp Ser Glu Pro Val Leu 1 5 10 <210> 26 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 26 Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe 1 5 10 <210> 27 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 27 Val Ile Thr Pro Gly Thr Asn Thr Ser Asn 1 5 10 <210> 28 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 28 Thr Val Cys Gly Pro Lys Lys Ser Thr Asn 1 5 10 <210> 29 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 29 Pro Ala Gln Glu Lys Asn Phe Thr Thr 1 5 <210> 30 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 30 Val Glu Gln Asp Lys Asn Thr Gln Glu 1 5 <210> 31 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 31 Asn Asn Leu Asp Ser Lys Val Gly Gly 1 5 <210> 32 <211> 42 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 32 agggacatcg ccgacaccac cgacgccgtg agggaccccc ag 42 <210> 33 <211> 42 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 33 gtgaggcaga tcgcccccgg ccagaccggc aagatcgccg ac 42 <210> 34 <211> 39 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 34 cagacccaga ccaacagccc caggagggcc aggagcgtg 39 <210> 35 <211> 39 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 35 ctgacccccg gcgacagcag cagcggctgg accgccggc 39 <210> 36 <211> 36 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 36 gtgtacgacc ccctgcagcc cgagctggac agcttc 36 <210> 37 <211> 36 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 37 atcctgcccg accccagcaa gcccagcaag aggagc 36 <210> 38 <211> 36 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 38 tacggcttcc agcccaccaa cggcgtgggc taccag 36 <210> 39 <211> 33 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 39 aagaaccaca ccagccccga cgtggacctg ggc 33 <210> 40 <211> 33 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 40 taccaggccg gcagcacccc ctgcaacggc gtg 33 <210> 41 <211> 33 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 41 agcggcacca acggcaccaa gaggttcgac aac 33 <210> 42 <211> 33 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 42 aggacccagc tgccccccgc ctacaccaac agc 33 <210> 43 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 43 ttcgacgagg acgacagcga gcccgtgctg 30 <210> 44 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 44 atctacaaga ccccccccat caaggacttc 30 <210> 45 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 45 gtgatcaccc ccggcaccaa caccagcaac 30 <210> 46 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 46 accgtgtgcg gccccaagaa gagcaccaac 30 <210> 47 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 47 cccgcccagg agaagaactt caccacc 27 <210> 48 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 48 gtggagcagg acaagaacac ccaggag 27 <210> 49 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 49 aacaacctgg acagcaaggt gggcggc 27 <210> 50 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 50 Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg 1 5 10 15 <210> 51 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 51 Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg 1 5 10 <210> 52 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 52 Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser Gly 1 5 10 15 <210> 53 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 53 Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr 1 5 10 <210> 54 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 54 Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly 1 5 10 15 Phe <210> 55 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 55 Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala 1 5 10 <210> 56 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 56 Val Ile Thr Pro Gly Thr Asn Thr Ser Asn 1 5 10 <210> 57 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 57 Thr Val Cys Gly Pro Lys Lys Ser Thr Asn 1 5 10 <210> 58 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 58 Pro Ala Gln Glu Lys Asn Phe Thr Thr 1 5 <210> 59 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 59 Val Glu Gln Asp Lys Asn Thr Gln Glu 1 5 <210> 60 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 60 Asn Asn Leu Asp Ser Lys Val Gly Gly 1 5 <210> 61 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 61 Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg 1 5 10 15 <210> 62 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 62 Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg 1 5 10 <210> 63 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 63 Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser Gly 1 5 10 15 <210> 64 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 64 Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr 1 5 10 <210> 65 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 65 Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly 1 5 10 15 Phe <210> 66 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 66 Gln Thr Gln Thr Asn Ser Pro Arg Arg Ala 1 5 10 <210> 67 <211> 3822 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 67 atgtttgtttt ttctgtttt attgccacta gtctctagtc atgtgttaa tcttacaacc 60 agaactcaat taccccctgc atacactaat tctttcacac gtggtgttta taccctgac 120 aaagttttca catcctcagttcattca actcaggact tgtctttca ttcttttcc 180 aatgttactt ggttccatgc tatacatgtc tctgggacca atggtactaa gaggtttgat 240 aaccctgtcc taccatta tgatgtgtt tattttgctt ccactgagaa gtctaacata 300 ataagaggct ggatttttgg tactacttta gattcgaaga cccagtccct acttattgtt 360 ataacgcta ctaatgttgt tattaaagtc tgtgaatttc aattttgtaa tgatccattt 420 ttgggtgtttt attaccacaaaacaaaa agttggatgg aaagtgagtt cagagtttat 480 tctagtgcga atattgcac ttttgaatat gtctctcagc cttttcttat ggaccttgaa 540 ggaaaacagg gtaatttca aaatcttagg gatttgtgt ttaagaat tgatggttat 600 tttaaaatat attctaagca cacgcctatt aatttagtgc gtgatctccc tcagggtttt 660 tcggctttag aaccattggt agatttgcca atagtatta acatcactag gtttcaact 720 ttacttgctt tacatagaag ttattgact cctggtgatt cttcttcagg ttggacagct 780 ggtgctgcag cttattatgt gggttatctt caacctagga cttttctatt aaatataat 840 gaaaatggaa ccattacaga tgctgtagac tgtgcacttg accctctc agaaacaaag 900 tgtacgttga aatccttcac tgtagaaaaa ggaatctatc aaacttctaa ctttagagtc 960 caaccaacag aatctattgt tagatttcct aatattacaa acttgtgccc ttttggtgaa 1020 gttttaacg ccaccagatt tgcatctgtt tatgcttgga acaggaagag aatcagcaac 1080 tgtgttgctg attattctgt cctatataat tccgcatcat tttccacttt taagtgttat 1140 ggagtgtctc ctactaaatt aaatgatctc tgctttacta atgtctatgc agattcattt 1200 gtaattagag gtgatgaagt cagacaaatc gctccagggc aaactggaaa gattgctgat 1260 tataattata aattaccaga tgattttaca ggctgcgtta tagcttggaa ttctaacaat 1320 cttgattcta aggttggtgg taattataat tacctgtata gattgtttag gaagtctaat 1380 ctcaaacctt ttgagagaga tatttcaact gaaatctatc aggccggtag cacaccttgt 1440 aatggtgttg aaggttttaa ttgttacttt cctttacaat catatggttt ccaacccact 1500 aatggtgttg gttaccaacc atacagagta gtagtacttt cttttgaact tctacatgca 1560 ccagcaactg tttgtggacc taaaaagtct actaatttgg ttaaaaacaa atgtgtcaat 1620 ttcaacttca atggtttaac aggcacaggt gttcttactg agtctaacaa aaagtttctg 1680 cctttccaac aatttggcag agacattgct gacactactg atgctgtccg tgatccacag 1740 acacttgaga ttcttgacat tacaccatgt tcttttggtg gtgtcagtgt tataacacca 1800 ggaacaaata cttctaacca ggttgctgtt ctttatcagg gtgttaactg cacagaagtc 1860 cctgttgcta ttcatgcaga tcaacttact cctacttggc gtgtttattc tacaggttct 1920 aatgtttttc aaacacgtgc aggctgttta ataggggctg aacatgtcaa caactcatat 1980 gagtgtgaca tacccattgg tgcaggtata tgcgctagtt atcagactca gactaattct 2040 ccttcgcggg caggtagtgt agctagtcaa tccatcattg cctacactat gtcacttggt 2100 gcagaaaatt cagttgctta ctctaataac tctattgcca tacccacaaa ttttactatt 2160 agtgttacca cagaaattct accagtgtct atgaccaaga catcagtaga ttgtacaatg 2220 tacatttgtg gtgattcaac tgaatgcagc aatcttttgt tgcaatatgg cagtttttgt 2280 acacaattaa accgtgcttt aactggaata gctgttgaac aagacaaaaa cacccaagaa 2340 gtttttgcac aagtcaaaca aatttacaaa acaccaccaa ttaaagattt tggtggtttt 2400 aatttttcac aaatattacc agatccatca aaaccaagca agaggtcacc tattgaagat 2460 ctacttttca acaaagtgac acttgcagat gctggcttca tcaaacaata tggtgattgc 2520 cttggtgata ttgctgctag agacctcatt tgtgcacaaa agtttaacgg ccttactgtt 2580 ttgccacctt tgctcacaga tgaaatgatt gctcaataca cttctgcact gttagcgggt 2640 acaatcactt ctggttggac ctttggtgca ggtcctgcat tacaaatacc atttcctatg 2700 caaatggctt ataggtttaa tggtattgga gttacacaga atgttctcta tgagaaccaa 2760 aaattgattg ccaaccaatt taatagtgct attggcaaaa ttcaagactc actttcttcc 2820 acaccaagtg cacttggaaa acttcaagat gtggtcaacc aaaatgcaca agctttaaac 2880 acgcttgtta aacaacttag ctccaatttt ggtgcaattt caagtgtttt aaatgatatc 2940 ctttcacgtc ttgacccacc tgaggctgaa gtgcaaattg ataggttgat cacaggcaga 3000 cttcaaagtt tgcagacata tgtgactcaa caattaatta gagctgcaga aatcagagct 3060 tctgctaatc ttgctgctac taaaatgtca gagtgtgtac ttggacaatc aaaaagagtt 3120 gatttttgtg gaaagggcta tcatcttatg tccttccctc agtcagcacc tcatggtgta 3180 gtcttcttgc atgtgactta tgtccctgca caagaaaga acttcacaac tgctcctgcc 3240 atttgtcatg atggaaaagc acactttcct cgtgaaggtg tctttgtttc aaatggcaca 3300 cactggtttg taacacaaag gaatttttat gaaccacaaa tcattactac agacaacaca 3360 tttgtgtctg gtaactgtga tgttgtaata ggaattgtca acaacacagt ttatgatcct 3420 ttgcaacctg aattagactc attcaaggag gagttagata aatattttaa gaatcataca 3480 tcaccagatg ttgatttagg tgacatctct ggcattaatg cttcagttgt aaacattcaa 3540 aaagaaattg accgcctcaa tgaggttgcc aagaatttaa atgaatctct catcgatctc 3600 caagaacttg gaaagtatga gcagtatata aaatggccat ggtacatttg gctaggtttt 3660 atagctggct tgattgccat agtaatggtg acaattatgc tttgctgtat gaccagttgc 3720 tgtagttgtc tcaagggctg ttgttcttgt ggatcctgct gcaaatttga tgaagacgac 3780 tctgagccag tgctcaaagg agtcaaatta cattacacat aa 3822 <210> 68 <211> 1273 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 68 Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val 1 5 10 15 Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe 20 25 30 Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu 35 40 45 His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp 50 55 60 Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp 65 70 75 80 Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu 85 90 95 Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser 100 105 110 Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile 115 120 125 Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr 130 135 140 Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr 145 150 155 160 Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu 165 170 175 Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe 180 185 190 Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr 195 200 205 Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu 210 215 220 Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr 225 230 235 240 Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser 245 250 255 Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro 260 265 270 Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala 275 280 285 Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys 290 295 300 Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val 305 310 315 320 Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys 325 330 335 Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala 340 345 350 Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu 355 360 365 Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro 370 375 380 Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe 385 390 395 400 Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly 405 410 415 Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys 420 425 430 Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn 435 440 445 Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe 450 455 460 Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys 465 470 475 480 Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly 485 490 495 Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val 500 505 510 Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys 515 520 525 Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn 530 535 540 Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu 545 550 555 560 Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val 565 570 575 Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe 580 585 590 Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val 595 600 605 Ala Val Leu Tyr Gln Gly Val Asn Cys Thr Glu Val Pro Val Ala Ile 610 615 620 His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser 625 630 635 640 Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val 645 650 655 Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala 660 665 670 Ser Tyr Gln Thr Gln Thr Asn Ser Pro Ser Arg Ala Gly Ser Val Ala 675 680 685 Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser 690 695 700 Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile 705 710 715 720 Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val 725 730 735 Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu 740 745 750 Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr 755 760 765 Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln 770 775 780 Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe 785 790 795 800 Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser 805 810 815 Pro Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly 820 825 830 Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp 835 840 845 Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu 850 855 860 Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly 865 870 875 880 Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Pro Ala Leu Gln Ile 885 890 895 Pro Phe Pro Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr 900 905 910 Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn 915 920 925 Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Pro Ser Ala 930 935 940 Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn 945 950 955 960 Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val 965 970 975 Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala Glu Val Gln 980 985 990 Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val 995 1000 1005 Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn 1010 1015 1020 Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys 1025 1030 1035 Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro 1040 1045 1050 Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val 1055 1060 1065 Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His 1070 1075 1080 Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn 1085 1090 1095 Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln 1100 1105 1110 Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val 1115 1120 1125 Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro 1130 1135 1140 Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn 1145 1150 1155 His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn 1160 1165 1170 Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu 1175 1180 1185 Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu 1190 1195 1200 Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu 1205 1210 1215 Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met 1220 1225 1230 Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys 1235 1240 1245 Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro 1250 1255 1260 Val Leu Lys Gly Val Lys Leu His Tyr Thr 1265 1270 <210> 69 <211> 816 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 69 gccaccatgt tcgtgtttct ggtgctgctg cctctggtgt ccagccagcg ggtgcagccc 60 accgaatcca tcgtgcggtt ccccaatatc accaatctgt gccccttcgg cgaggtgttc 120 aatgccacca gattcgcctc tgtgtacgcc tggaaccgga agcggatcag caattgcgtg 180 gccgactact ccgtgctgta caactccgcc agcttcagca ccttcaagtg ctacggcgtg 240 tcccctacca agctgaacga cctgtgcttc acaaacgtgt acgccgacag cttcgtgatc 300 cggggagatg aagtgcggca gattgcccct ggacagacag gcaagatcgc cgactacaac 360 tacaagctgc ccgacgactt caccggctgt gtgattgcct ggaacagcaa caacctggac 420 tccaaagtcg gcggcaacta caattacctg taccggctgt tccggaagtc caatctgaag 480 cccttcgagc gggacatctc caccgagatc tatcaggccg gcagcacccc ttgtaacggc 540 gtggaaggct tcaactgcta cttcccactg cagtcctacg gctttcagcc cacaaatggc 600 gtgggctatc agccctacag agtggtggtg ctgagcttcg aactgctgca tgcccctgcc 660 acagtgtgcg gccctaagaa aagcaccaat ctcgtgaaga acaaatgcgt gaacttcgga 720 tccggaggtg gatacatccc ggaggccct agggacggtc aagcttacgt gagaaaggac 780 ggcgaatggg ttctgctgtc gaccttctg ggataa 816 <210> 70 <211> 271 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 70 Ala Thr Met Phe Val Phe Val Leu Val Leu Leu Pro Leu Val Ser Ser Gln 1 5 10 15 Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn 20 25 30 Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val 35 40 45 Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser 50 55 60 Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val 65 70 75 80 Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp 85 90 95 Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln 100 105 110 Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr 115 120 125 Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly 130 135 140 Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys 145 150 155 160 Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr 165 170 175 Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser 180 185 190 Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val 195 200 205 Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly 210 215 220 Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Gly 225 230 235 240 Ser Gly Gly Gly Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr 245 250 255 Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu Gly 260 265 270 <210> 71 <211> 3822 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 71 atgtttgttt ttcttgtttt attgccacta gtctctagtc agtgtgttaa tcttacaacc 60 agaactcaat taccccctgc atacactaat tctttcacac gtggtgttta ttaccctgac 120 aaagttttca gatcctcagt tttacattca actcaggact tgttcttacc tttcttttcc 180 aatgttactt ggttccatgc tatacatgtc tctgggacca atggtactaa gaggtttgat 240 aaccctgtcc taccatttaa tgatggtgtt tattttgctt ccactgagaa gtctaacata 300 ataagaggct ggatttttgg tactacttta gattcgaaga cccagtccct acttattgtt 360 aataacgcta ctaatgttgt tattaaagtc tgtgaatttc aattttgtaa tgatccattt 420 ttgggtgttt attaccacaa aaacaacaaa agttggatgg aaagtgagtt cagagtttat 480 tctagtgcga atattgcac ttttgaatat gtctctcagc cttttcttat ggaccttgaa 540 ggaaaacagg gtaatttca aaatcttagg gatttgtgt ttaagaat tgatggttat 600 tttaaaatat attctaagca cacgcctatt aatttagtgc gtgatctccc tcagggtttt 660 tcggctttag aaccattggt agatttgcca atagtatta acatcactag gtttcaact 720 ttacttgctt tacatagaag tatttgact cctggtgat cttctcagg ttggacagct 780 ggtgctgcag cttattatgt gggttatctt caactagga ctttctatt aaaatataat 840 gaaaatggaa ccattacaga tgctgtagac tgtgcacttg accctctc agaaacaaag 900 tgtacgttga atccttcac tgtagaaaaa ggaatctatc aaacttctaa ctttagagtc 960 caccacag aatctattgt tagattcct atattacaa acttgtgccc ttttggtgaa 1020 gtttttaacg ccaccagatt tgcatctgtt tatgcttgga acaggaagg atcagcaac 1080 tgtgttgctg attattctgt cctatataat tccgcatcat tttccactt taagtgttat 1140 ggagtgtctc ctactaaatt aaatgatctc tgcttacta atgtctatgc agattcattt 1200 gtaattagag gtgatgaagt cagacaaatc gctccagggc aaactggaaa gattgctgat 1260 tataattata aattaccaga tgattttaca ggctgcgtta tagcttggaa ttctaacaat 1320 cttgattcta aggttggtgg taattataat tacctgtata gattgtttag gaagtctaat 1380 ctcaaacctt ttgagagaga tatttcaact gaaatctatc aggccggtag cacaccttgt 1440 aatggtgttg aaggttttaa ttgttacttt cctttacaat catatggttt ccaacccact 1500 tatggtgttg gttaccaacc atacagagta gtagtacttt cttttgaact tctacatgca 1560 ccagcaactg tttgtggacc taaaaagtct actaatttgg ttaaaaacaa atgtgtcaat 1620 ttcaacttca atggtttaac aggcacaggt gttcttactg agtctaacaa aaagtttctg 1680 cctttccaac aatttggcag agacattgat gacactactg atgctgtccg tgatccacag 1740 acacttgaga ttcttgacat tacaccatgt tctttttggtg gtgtcagtgt tataacacca 1800 ggaacaaata cttctaacca ggttgctgtt ctttatcagg gtgttaactg cacagaagtc 1860 cctgttgcta ttcatgcaga tcaacttact cctacttggc gtgttattc tacaggttct 1920 aatgtttttc aaacacgtgc aggctgttta ataggggctg aacatgtcaa caactcatat 1980 gagtgtgaca tacccattgg tgcaggtata tgcgctagtt atcagactca gactaattct 2040 cattcgcggg caggtagtgt agctagtcaa tccatcattg cctacactat gtcacttggt 2100 gcagaaaatt cagttgctta ctctaataac tctattgcca tacccataaa ttttactatt 2160 agtgttacca cagaaattct accagtgtct atgaccaaga catcagtaga ttgtacaatg 2220 tacatttgtg gtgattcaac tgaatgcagc aatcttttgt tgcaatatgg cagtttttgt 2280 acacaattaa accgtgcttt aactggaata gctgttgaac aagacaaaaa cacccaagaa 2340 gtttttgcac aagtcaaaca aatttacaaa acaccaccaa ttaaagattt tggtggtttt 2400 aatttttcac aaatattacc agatccatca aaaccaagca agaggtcacc tattgaagat 2460 ctacttttca acaaagtgac acttgcagat gctggcttca tcaaacaata tggtgattgc 2520 cttggtgata ttgctgctag agacctcatt tgtgcacaaa agtttaacgg ccttactgtt 2580 ttgccacctt tgctcacaga tgaaatgatt gctcaataca cttctgcact gttagcgggt 2640 acaatcactt ctggttggac ctttggtgca ggtcctgcat tacaaatacc atttcctatg 2700 caaatggctt ataggtttaa tggtattgga gttacacaga atgttctcta tgagaaccaa 2760 aaattgattg ccaaccaatt taatagtgct attggcaaaa ttcaagactc actttcttcc 2820 acaccaagtg cacttggaaa acttcaagat gtggtcaacc aaaatgcaca agctttaaac 2880 acgcttgtta aacaacttag ctccaatttt ggtgcaattt caagtgtttt aaatgatatc 2940 cttgcacgtc ttgacccacc tgaggctgaa gtgcaaattg ataggttgat cacaggcaga 3000 cttcaaagtt tgcagacata tgtgactcaa caattaatta gagctgcaga aatcagagct 3060 tctgctaatc ttgctgctac taaaatgtca gagtgtgtac ttggacaatc aaaaagagtt 3120 gatttttgtg gaaagggcta tcatcttatg tccttccctc agtcagcacc tcatggtgta 3180 gtcttcttgc atgtgactta tgtccctgca caagaaaaga acttcacaac tgctcctgcc 3240 atttgtcatg atggaaaagc acactttcct cgtgaaggtg tctttgtttc aaatggcaca 3300 cactggtttg taacacaaag gaatttttat gaaccacaaa tcattactac acacaacaca 3360 tttgtgtctg gtaactgtga tgttgtaata ggaattgtca acaacacagt ttatgatcct 3420 ttgcaacctg aattagactc attcaaggag gagttagata aatattttaa gaatcataca 3480 tcaccagatg ttgatttagg tgacatctct ggcattaatg cttcagttgt aaacattcaa 3540 aaagaaattg accgcctcaa tgaggttgcc aagaatttaa atgaatctct catcgatctc 3600 caagaacttg gaaagtatga gcagtatata aaatggccat ggtacatttg gctaggtttt 3660 atagctggct tgattgccat agtaatggtg acaattatgc tttgctgtat gaccagttgc 3720 tgtagttgtc tcaagggctg ttgttcttgt ggatcctgct gcaaatttga tgaagacgac 3780 tctgagccag tgctcaaagg agtcaaatta cattacacat aa 3822 <210> 72 <211> 1273 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 72 Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val 1 5 10 15 Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe 20 25 30 Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu 35 40 45 His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp 50 55 60 Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp 65 70 75 80 Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu 85 90 95 Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser 100 105 110 Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile 115 120 125 Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr 130 135 140 Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr 145 150 155 160 Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu 165 170 175 Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe 180 185 190 Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr 195 200 205 Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu 210 215 220 Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr 225 230 235 240 Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser 245 250 255 Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro 260 265 270 Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala 275 280 285 Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys 290 295 300 Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val 305 310 315 320 Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys 325 330 335 Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala 340 345 350 Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu 355 360 365 Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro 370 375 380 Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe 385 390 395 400 Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly 405 410 415 Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys 420 425 430 Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn 435 440 445 Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe 450 455 460 Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys 465 470 475 480 Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly 485 490 495 Phe Gln Pro Thr Tyr Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val 500 505 510 Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys 515 520 525 Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn 530 535 540 Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu 545 550 555 560 Pro Phe Gln Gln Phe Gly Arg Asp Ile Asp Asp Thr Thr Asp Ala Val 565 570 575 Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe 580 585 590 Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val 595 600 605 Ala Val Leu Tyr Gln Gly Val Asn Cys Thr Glu Val Pro Val Ala Ile 610 615 620 His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser 625 630 635 640 Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val 645 650 655 Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala 660 665 670 Ser Tyr Gln Thr Gln Thr Asn Ser His Ser Arg Ala Gly Ser Val Ala 675 680 685 Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser 690 695 700 Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Ile Asn Phe Thr Ile 705 710 715 720 Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val 725 730 735 Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu 740 745 750 Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr 755 760 765 Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln 770 775 780 Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe 785 790 795 800 Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser 805 810 815 Pro Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly 820 825 830 Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp 835 840 845 Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu 850 855 860 Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly 865 870 875 880 Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Pro Ala Leu Gln Ile 885 890 895 Pro Phe Pro Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr 900 905 910 Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn 915 920 925 Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Pro Ser Ala 930 935 940 Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn 945 950 955 960 Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val 965 970 975 Leu Asn Asp Ile Leu Ala Arg Leu Asp Pro Pro Glu Ala Glu Val Gln 980 985 990 Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val 995 1000 1005 Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn 1010 1015 1020 Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys 1025 1030 1035 Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro 1040 1045 1050 Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val 1055 1060 1065 Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His 1070 1075 1080 Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn 1085 1090 1095 Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln 1100 1105 1110 Ile Ile Thr Thr His Asn Thr Phe Val Ser Gly Asn Cys Asp Val 1115 1120 1125 Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro 1130 1135 1140 Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn 1145 1150 1155 His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn 1160 1165 1170 Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu 1175 1180 1185 Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu 1190 1195 1200 Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu 1205 1210 1215 Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met 1220 1225 1230 Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys 1235 1240 1245 Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro 1250 1255 1260 Val Leu Lys Gly Val Lys Leu His Tyr Thr 1265 1270 <210> 73 <211> 3813 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 73 atgtttgttt ttcttgtttt attgccacta gtctctagtc agtgtgttaa tcttacaacc 60 agaactcaat taccccctgc atacactaat tctttcacac gtggtgttta ttaccctgac 120 aaagttttca gatcctcagt tttacattca actcaggact tgttcttacc tttcttttcc 180 aatgttactt ggttccatgc tatatctggg accaatggta ctaagaggtt tgataaccct 240 gtcctaccat ttaatgatgg tgtttatttt gcttccactg agaagtctaa cataataaga 300 ggctggattt ttggtactac tttagattcg aagacccagt ccctacttat tgttaataac 360 gctactaatg ttgttattaa agtctgtgaa tttcaatttt gtaatgatcc atttttgggt 420 gtttaccaca aaaacaacaa aagttggatg gaaagtgagt tcagagttta ttctagtgcg 480 aataattgca cttttgaata tgtctctcag ccttttctta tggaccttga aggaaaacag 540 ggtaatttca aaaatcttag ggaatttgtg tttaagaata ttgatggtta ttttaaaata 600 tattctaagc acacgcctat taatttagtg cgtgatctcc ctcagggttt ttcggcttta 660 gaaccattgg tagatttgcc aataggtatt aacatcacta ggtttcaaac tttacttgct 720 ttacatagaa gttattgac tcctggtgat tcttcttcag gttggacagc tggtgctgca 780 gcttattatg tgggttatct tcaacctagg actttctat taaaatataa tgaaaatgga 840 accattacag atgctgtaga ctgtgcactt gaccctctct cagaaacaaa gtgtacgttg 900 aaatccttca ctgtagaaaa aggaatctat caaacttcta actttagagt ccaaccaaca 960 gaatctattg ttagatttcc taatattaca aacttgtgcc cttttggtga agtttttaac 1020 gccaccagat ttgcatctgt ttatgcttgg aacaggaaga gaatcagcaa ctgtgttgct 1080 gattattctg tcctatataa ttccgcatca ttttccactt ttaagtgtta tggagtgtct 1140 cctactaaat taaatgatct ctgctttact aatgtctatg cagattcatt tgtaattaga 1200 ggtgatgaag tcagacaaat cgctccaggg caaactggaa agattgctga ttataattat 1260 aaattaccag atgattttac aggctgcgtt atagcttgga attctaacaa tcttgattct 1320 aaggttggtg gtaattataa ttacctgtat agattgttta ggaagtctaa tctcaaacct 1380 tttgagagag atattcaac tgaaatctat caggccggta gcacaccttg taatggtgtt 1440 gaaggtttta attgttactt tcctttacaa tcatatggtt tccaacccac ttatggtgtt 1500 ggttaccaac catacagagt agtagtactt tcttttgaac ttctacatgc accagcaact 1560 gttgtggac ctaaaaagtc tactaatttg gttaaaaaca aatgtgtcaa tttcaacttc 1620 aatggtttaa caggcacagg tgttcttact gagtctaaca aaaagtttct gcctttccaa 1680 caatttggca gagacattga tgacactact gatgctgtcc gtgatccaca gacacttgag 1740 attcttgaca ttacaccatg ttctttggt ggtgtcagtg ttataacacc aggaacaaat 1800 acttctaacc aggttgctgt tctttatcag ggtgttaact gcacagaagt ccctgttgct 1860 attcatgcag atcaacttac tcctacttgg cgtgtttatt ctacaggttc taatgttttt 1920 1980 atacccattg gtgcaggtat atgcgctagt tatcagactc agactaattc tcattcgcgg 2040 gcaggtagtg tagctagtca atccatcatt gcctacacta tgtcacttgg tgcagaaaat 2100 tcagttgctt actctaataa ctctattgcc atacccataa attttactat tagtgttacc 2160 agaaattc taccagtgtc tatgaccaag acatcagtag attgtacaat gtacatttgt 2220 ggtgattcaa ctgaatgcag caatcttttg ttgcaatatg gcagtttttg tacaaatta 2280 aaccgtgctt taactggaat agctgttgaa caagacaaaa acacccaaga agttttgca 2340 caagtcaac aaatttacaa aacaccacca attaaagatt ttggtggttt taatttttca 2400 caaatattac cagatccatc aaaaccaagc aagaggtcac ctattgaaga tctacttttc 2460 aaaaagtga cacttgcaga tgctggcttc atcaaacaat atggtgattg ccttggtgat 2520 attgctgcta gagacctcat ttgtgcacaa aagtttaacg gccttactgt tttgccacct 2580 ttgctcacag atgaaatgat tgctcaatac acttctgcac tgttagcggg tacaatcact 2640 tctggttgga cctttggtgc aggtcctgca ttacaaatac catttcctat gcaaatggct 2700 tataggttta atggtattgg agttacacag aatgttctct atgagaacca aaaattgatt 2760 gccaaccaat ttaatagtgc tattggcaaa attcaagact cactttcttc cacaccaagt 2820 gcacttggaa aacttcaaga tgtggtcaac caaaatgcac aagctttaaa cacgcttgtt 2880 aaacaactta gctccaattt tggtgcaatt tcaagtgttt taaatgatat ccttgcacgt 2940 cttgacccac ctgaggctga agtgcaaatt gataggttga tcacaggcag acttcaaagt 3000 ttgcagacat atgtgactca acaattaatt agagctgcag aaatcagagc ttctgctaat 3060 cttgctgcta ctaaaatgtc agagtgtgta cttggacaat caaaaagagt tgatttttgt 3120 ggaaagggct atcatcttat gtccttccct cagtcagcac ctcatggtgt agtcttcttg 3180 catgtgactt atgtccctgc acaagaaaag aacttcacaa ctgctcctgc catttgtcat 3240 gatggaaaag cacactttcc tcgtgaaggt gtctttgttt caaatggcac acactggttt 3300 gtaacacaaa ggaattttta tgaaccacaa atcattacta cacacaacac atttgtgtct 3360 ggtaactgtg atgttgtaat aggaattgtc aacaacacag tttatgatcc tttgcaacct 3420 gaattagact cattcaagga ggagttagat aaatatttta agaatcatac atcaccagat 3480 gttgatttag gtgacatctc tggcattaat gcttcagttg taaacattca aaaagaaatt 3540 gaccgcctca atgaggttgc caagaattta aatgaatctc tcatcgatct ccaagaactt 3600 ggaaagtatg agcagtatat aaaatggcca tggtacattt ggctaggttt tatagctggc 3660 ttgattgcca tagtaatggt gacaattatg ctttgctgta tgaccagttg ctgtagttgt 3720 ctcaagggct gttgttcttg tggatcctgc tgcaaatttg atgaagacga ctctgagcca 3780 gtgctcaaag gagtcaaatt acattacaca taa 3813 <210> 74 <211> 1270 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 74 Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val 1 5 10 15 Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe 20 25 30 Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu 35 40 45 His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp 50 55 60 Phe His Ala Ile Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp Asn Pro 65 70 75 80 Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu Lys Ser 85 90 95 Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser Lys Thr 100 105 110 Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile Lys Val 115 120 125 Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr His Lys 130 135 140 Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr Ser Ser Ala 145 150 155 160 Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu Met Asp Leu 165 170 175 Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe Val Phe Lys 180 185 190 Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr Pro Ile Asn 195 200 205 Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu Pro Leu Val 210 215 220 Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr Leu Leu Ala 225 230 235 240 Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser Gly Trp Thr 245 250 255 Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro Arg Thr Phe 260 265 270 Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala Val Asp Cys 275 280 285 Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys Ser Phe Thr 290 295 300 Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val Gln Pro Thr 305 310 315 320 Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly 325 330 335 Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg 340 345 350 Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser 355 360 365 Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu 370 375 380 Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg 385 390 395 400 Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala 405 410 415 Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala 420 425 430 Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr 435 440 445 Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp 450 455 460 Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val 465 470 475 480 Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro 485 490 495 Thr Tyr Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe 500 505 510 Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Thr 515 520 525 Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn Gly Leu Thr 530 535 540 Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu Pro Phe Gln 545 550 555 560 Gln Phe Gly Arg Asp Ile Asp Asp Thr Thr Asp Ala Val Arg Asp Pro 565 570 575 Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe Gly Gly Val 580 585 590 Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val Ala Val Leu 595 600 605 Tyr Gln Gly Val Asn Cys Thr Glu Val Pro Val Ala Ile His Ala Asp 610 615 620 Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser Asn Val Phe 625 630 635 640 Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val Asn Asn Ser 645 650 655 Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala Ser Tyr Gln 660 665 670 Thr Gln Thr Asn Ser His Ser Arg Ala Gly Ser Val Ala Ser Gln Ser 675 680 685 Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser Val Ala Tyr 690 695 700 Ser Asn Asn Ser Ile Ala Ile Pro Ile Asn Phe Thr Ile Ser Val Thr 705 710 715 720 Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val Asp Cys Thr 725 730 735 Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu Leu Leu Gln 740 745 750 Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr Gly Ile Ala 755 760 765 Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln Val Lys Gln 770 775 780 Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe Asn Phe Ser 785 790 795 800 Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser Pro Ile Glu 805 810 815 Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly Phe Ile Lys 820 825 830 Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys 835 840 845 Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp 850 855 860 Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr 865 870 875 880 Ser Gly Trp Thr Phe Gly Ala Gly Pro Ala Leu Gln Ile Pro Phe Pro 885 890 895 Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val 900 905 910 Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile 915 920 925 Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Pro Ser Ala Leu Gly Lys 930 935 940 Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val 945 950 955 960 Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp 965 970 975 Ile Leu Ala Arg Leu Asp Pro Pro Glu Ala Glu Val Gln Ile Asp Arg 980 985 990 Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln 995 1000 1005 Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala 1010 1015 1020 Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys Arg Val Asp 1025 1030 1035 Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro Gln Ser Ala 1040 1045 1050 Pro His Gly Val Val Phe Leu His Val Thr Tyr Val Pro Ala Gln 1055 1060 1065 Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp Gly Lys 1070 1075 1080 Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly Thr His 1085 1090 1095 Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr 1100 1105 1110 Thr His Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly 1115 1120 1125 Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp 1130 1135 1140 Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser 1145 1150 1155 Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala Ser Val 1160 1165 1170 Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu Val Ala Lys 1175 1180 1185 Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr 1190 1195 1200 Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu Gly Phe Ile 1205 1210 1215 Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met Leu Cys Cys 1220 1225 1230 Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys Ser Cys Gly 1235 1240 1245 Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro Val Leu Lys 1250 1255 1260 Gly Val Lys Leu His Tyr Thr 1265 1270 <210> 75 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 75 Lys His Lys His Lys His Lys His Lys 1 5 <210> 76 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 76 His Lys His Lys His Lys His Lys His Lys 1 5 10 <210> 77 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 77 Lys His Lys His Lys His Lys His Lys His 1 5 10 <210> 78 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 78 His Lys His Lys His Lys His Lys His Lys His 1 5 10 <210> 79 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 79 Lys His Lys His His Lys His His Lys His His Lys His His Lys His 1 5 10 15 His Lys His Lys 20 <210> 80 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 80 Lys His His His Lys His His His Lys His His His Lys His His His 1 5 10 15 Lys <210> 81 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 81 Lys His His His Lys His His His Lys His His His His Lys His His 1 5 10 15 His Lys <210> 82 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <220> <221> MOD_RES <222> (1)..(1) <223> D-amino acid <220> <221> MOD_RES <222> (5)..(5) <223> D-amino acid <220> <221> MOD_RES <222> (9)..(9) <223> D-amino acid <220> <221> MOD_RES <222> (14)..(14) <223> D-amino acid <220> <221> MOD_RES <222> (18)..(18) <223> D-amino acid <400> 82 Lys His His His Lys His His His Lys His His His His Lys His His 1 5 10 15 His Lys <210> 83 <211> 19 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 83 His Lys His His His Lys His His His Lys His His His His Lys His 1 5 10 15 His His Lys <210> 84 <211> 20 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 84 His His Lys His His His Lys His His His Lys His His His His Lys 1 5 10 15 His His His Lys 20 <210> 85 <211> 21 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 85 Lys His His His His Lys His His His His Lys His His His His Lys 1 5 10 15 His His His His Lys 20 <210> 86 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 86 Lys His His His Lys His His His Lys His His His Lys His His His 1 5 10 15 His Lys <210> 87 <211> 18 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 87 Lys His His His Lys His His His His Lys His His His Lys His His 1 5 10 15 His Lys <210> 88 <211> 19 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 88 Lys His His His Lys His His His His Lys His His His Lys His His 1 5 10 15 His His Lys <210> 89 <211> 3822 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polynucleotide <400> 89 atgtttgttt ttcttgtttt attgccacta gtctctagtc agtgtgttaa tcttacaacc 60 agaactcaat taccccctgc atacactaat tctttcacac gtggtgttta ttaccctgac 120 aaagttttca gatcctcagt tttacattca actcaggact tgttcttacc tttcttttcc 180 aatgttactt ggttccatgc tatacatgtc tctgggacca atggtactaa gaggtttgat 240 aaccctgtcc taccatttaa tgatggtgtt tattttgctt ccactgagaa gtctaacata 300 ataagaggct ggatttttgg tactacttta gattcgaaga cccagtccct acttattgtt 360 ataacgcta ctaatgttgt tattaaagtc tgtgaatttc aattttgtaa tgatccattt 420 ttgggtgtttt attaccacaaaacaaaa agttggatgg aaagtgagtt cagagtttat 480 tctagtgcga atattgcac ttttgaatat gtctctcagc cttttcttat ggaccttgaa 540 ggaaaacagg gtaatttca aaatcttagg gatttgtgt ttaagaat tgatggttat 600 tttaaaatat attctaagca cacgcctatt aatttagtgc gtgatctccc tcagggtttt 660 tcggctttag aaccattggt agatttgcca atagtatta acatcactag gtttcaact 720 ttacttgctt tacatagaag tatttgact cctggtgat cttctcagg ttggacagct 780 ggtgctgcag cttattatgt gggttatctt caactagga ctttctatt aaaatataat 840 gaaaatggaa ccattacaga tgctgtagac tgtgcacttg accctctc agaaacaaag 900 tgtacgttga atccttcac tgtagaaaaa ggaatctatc aaacttctaa ctttagagtc 960 caccacag aatctattgt tagattcct atattacaa acttgtgccc ttttggtgaa 1020 gtttttaacg ccaccagatt tgcatctgtt tatgcttgga acaggaagg atcagcaac 1080 tgtgttgctg attattctgt cctatataat tccgcatcat tttccacttt taagtgttat 1140 ggagtgtctc ctactaaatt aaatgatctc tgctttacta atgtctatgc agattcattt 1200 gtaattagag gtgatgaagt cagacaaatc gctccagggc aaactggaaa gattgctgat 1260 tataattata aattaccaga tgattttaca ggctgcgtta tagcttggaa ttctaacaat 1320 cttgattcta aggttggtgg taattataat tacctgtata gattgtttag gaagtctaat 1380 ctcaaacctt ttgagagaga tatttcaact gaaatctatc aggccggtag cacaccttgt 1440 aatggtgttg aaggttttaa ttgttacttt cctttacaat catatggttt ccaacccact 1500 aatggtgttg gttaccaacc atacagagta gtagtacttt cttttgaact tctacatgca 1560 ccagcaactg tttgtggacc taaaaagtct actaatttgg ttaaaaacaa atgtgtcaat 1620 ttcaacttca atggtttaac aggcacaggt gttcttactg agtctaacaa aaagtttctg 1680 cctttccaac aatttggcag agacattgct gacactactg atgctgtccg tgatccacag 1740 acacttgaga ttcttgacat tacaccatgt tctttttggtg gtgtcagtgt tataacacca 1800 ggaacaaata cttctaacca ggttgctgtt ctttatcagg gtgttaactg cacagaagtc 1860 cctgttgcta ttcatgcaga tcaacttact cctacttggc gtgtttattc tacaggttct 1920 aatgtttttc aaacacgtgc aggctgttta ataggggctg aacatgtcaa caactcatat 1980 gagtgtgaca tacccattgg tgcaggtata tgcgctagtt atcagactca gactaattct 2040 ccttcgcggg caggtagtgt agctagtcaa tccatcattg cctacactat gtcacttggt 2100 gcagaaaatt cagttgctta ctctaataac tctattgcca tacccacaaa ttttactatt 2160 agtgttacca cagaaattct accagtgtct atgaccaaga catcagtaga ttgtacaatg 2220 tacatttgtg gtgattcaac tgaatgcagc aatcttttgt tgcaatatgg cagtttttgt 2280 acacaattaa accgtgcttt aactggaata gctgttgaac aagacaaaaa cacccaagaa 2340 gtttttgcac aagtcaaaca aatttacaaa acaccaccaa ttaaagattt tggtggtttt 2400 aatttttcac aaatattacc agatccatca aaaccaagca agaggtcatt tattgaagat 2460 ctacttttca acaaagtgac acttgcagat gctggcttca tcaaacaata tggtgattgc 2520 cttggtgata ttgctgctag agacctcatt tgtgcacaaa agtttaacgg ccttactgtt 2580 ttgccacctt tgctcacaga tgaaatgatt gctcaataca cttctgcact gttagcgggt 2640 acaatcactt ctggttggac ctttggtgca ggtgctgcat tacaaatacc atttgctatg 2700 caaatggctt ataggtttaa tggtattgga gttacacaga atgttctcta tgagaaccaa 2760 aaattgattg ccaaccaatt taatagtgct attggcaaaa ttcaagactc actttcttcc 2820 acagcaagtg cacttggaaa acttcaagat gtggtcaacc aaaatgcaca agctttaaac 2880 acgcttgtta aacaacttag ctccaatttt ggtgcaattt caagtgtttt aaatgatatc 2940 ctttcacgtc ttgacccacc tgaggctgaa gtgcaaattg ataggttgat cacaggcaga 3000 cttcaaagtt tgcagacata tgtgactcaa caattaatta gagctgcaga aatcagagct 3060 tctgctaatc ttgctgctac taaaatgtca gagtgtgtac ttggacaatc aaaaagagtt 3120 gatttttgtg gaaagggcta tcatcttatg tccttccctc agtcagcacc tcatggtgta 3180 gtcttcttgc atgtgactta tgtccctgca caagaaaaga acttcacaac tgctcctgcc 3240 atttgtcatg atggaaaagc acactttcct cgtgaaggtg tctttgtttc aaatggcaca 3300 cactggtttg taacacaaag gaatttttat gaaccacaaa tcattactac agacaacaca 3360 tttgtgtctg gtaactgtga tgttgtaata ggaattgtca acaacacagt ttatgatcct 3420 ttgcaacctg aattagactc attcaaggag gagttagata aatattttaa gaatcataca 3480 tcaccagatg ttgatttagg tgacatctct ggcattaatg cttcagttgt aaacattcaa 3540 aaagaaattg accgcctcaa tgaggttgcc aagaatttaa atgaatctct catcgatctc 3600 caagaacttg gaaagtatga gcagtatata aaatggccat ggtacatttg gctaggtttt 3660 atagctggct tgattgccat agtaatggtg acaattatgc tttgctgtat gaccagttgc 3720 tgtagttgtc tcaagggctg ttgttcttgt ggatcctgct gcaaatttga tgaagacgac 3780 tctgagccag tgctcaaagg agtcaaatta cattacacat aa 3822 <210> 90 <211> 1273 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 90 Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val 1 5 10 15 Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe 20 25 30 Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu 35 40 45 His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp 50 55 60 Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp 65 70 75 80 Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu 85 90 95 Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser 100 105 110 Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile 115 120 125 Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr 130 135 140 Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr 145 150 155 160 Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu 165 170 175 Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe 180 185 190 Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr 195 200 205 Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu 210 215 220 Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr 225 230 235 240 Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser 245 250 255 Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro 260 265 270 Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala 275 280 285 Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys 290 295 300 Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val 305 310 315 320 Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys 325 330 335 Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala 340 345 350 Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu 355 360 365 Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro 370 375 380 Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe 385 390 395 400 Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly 405 410 415 Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys 420 425 430 Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn 435 440 445 Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe 450 455 460 Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys 465 470 475 480 Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly 485 490 495 Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val 500 505 510 Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys 515 520 525 Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn 530 535 540 Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu 545 550 555 560 Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val 565 570 575 Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe 580 585 590 Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val 595 600 605 Ala Val Leu Tyr Gln Gly Val Asn Cys Thr Glu Val Pro Val Ala Ile 610 615 620 His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser 625 630 635 640 Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val 645 650 655 Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala 660 665 670 Ser Tyr Gln Thr Gln Thr Asn Ser Pro Ser Arg Ala Gly Ser Val Ala 675 680 685 Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser 690 695 700 Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile 705 710 715 720 Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val 725 730 735 Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu 740 745 750 Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr 755 760 765 Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln 770 775 780 Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe 785 790 795 800 Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser 805 810 815 Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly 820 825 830 Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp 835 840 845 Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu 850 855 860 Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly 865 870 875 880 Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile 885 890 895 Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr 900 905 910 Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn 915 920 925 Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala 930 935 940 Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn 945 950 955 960 Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val 965 970 975 Leu Asn Asp Ile Leu Ser Arg Leu Asp Pro Pro Glu Ala Glu Val Gln 980 985 990 Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val 995 1000 1005 Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn 1010 1015 1020 Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys 1025 1030 1035 Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro 1040 1045 1050 Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val 1055 1060 1065 Pro Ala Gln Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His 1070 1075 1080 Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn 1085 1090 1095 Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln 1100 1105 1110 Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val 1115 1120 1125 Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro 1130 1135 1140 Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn 1145 1150 1155 His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn 1160 1165 1170 Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu 1175 1180 1185 Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu 1190 1195 1200 Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Trp Tyr Ile Trp Leu 1205 1210 1215 Gly Phe Ile Ala Gly Leu Ile Ala Ile Val Met Val Thr Ile Met 1220 1225 1230 Leu Cys Cys Met Thr Ser Cys Cys Ser Cys Leu Lys Gly Cys Cys 1235 1240 1245 Ser Cys Gly Ser Cys Cys Lys Phe Asp Glu Asp Asp Ser Glu Pro 1250 1255 1260 Val Leu Lys Gly Val Lys Leu His Tyr Thr 1265 1270 <210> 91 <211> 36 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 91 gccatccacg tgagcggcac caacggcacc aagagg 36 <210> 92 <211> 45 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 92 gccctgcaca ggagctacct gacccccggc gacagcagca gcggc 45 <210> 93 <211> 33 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 93 aacaacctgg acagcaaggt gggcggcaac tac 33 <210> 94 <211> 51 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 94 accgagatct accaggccgg cagcaccccc tgcaacggcg tggagggctt c 51 <210> 95 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 95 cagacccaga ccaacagccc caggagggcc 30 <210> 96 <211> 30 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 96 gugaucaccc ccggcaccaa caccagcaac 30 <210> 97 <211> 30 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 97 accgugugcg gccccaagaa gagcaccaac 30 <210> 98 <211> 27 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 98 cccgcccagg agaagaacuu caccacc 27 <210> 99 <211> 27 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 99 guggagcagg acaagaacac ccaggag 27 <210> 100 <211> 27 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 100 aacaaccugg acagcaaggu gggcggc 27 <210> 101 <211> 45 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 101 uacuaccaca agaacaacaa gagcuggaug gagagcgagu ucagg 45 <210> 102 <211> 36 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 102 gccauccacg ugagcggcac caacggcacc aagagg 36 <210> 103 <211> 45 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 103 gcccugcaca ggagcuaccu gacccccggc gacagcagca gcggc 45 <210> 104 <211> 33 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 104 aacaaccugg acagcaaggu gggcggcaac uac 33 <210> 105 <211> 51 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 105 accgagaucu accaggccgg cagcaccccc ugcaacggcg uggagggcuu c 51 <210> 106 <211> 30 <212> RNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 106 cagacccaga ccaacagccc caggagggcc 30 <210> 107 <211> 4 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 107 His His His Lys 1 <210> 108 <211> 45 <212> DNA <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic oligonucleotide <400> 108 tactaccaca agaacaacaa gagctggatg gagagcgagt tcagg 45

Claims

1. A composition comprising an effective amount of messenger ribonucleic acid (mRNA) formulated in a pharmaceutically acceptable carrier, the messenger ribonucleic acid comprising an open reading frame (ORF) encoding a SARS-CoV-2 protein with the nucleotide sequence SEQ ID NO: 89, wherein the mRNA and the carrier form polymer nanoparticles, and the carrier comprises a histidine-lysine copolymer (HKP), wherein the HKP has the structural formula shown in Formula I. (Formula I) Where K represents L-lysine, and each of R1, R2, R3 and R4 is identical and selected from the amino acid sequence shown in SEQ ID NO: 81 or 82.

2. The composition according to claim 1, wherein the SARS-CoV-2 protein is a spike (S) protein or a spike (S) subunit.

3. The composition according to claim 2, wherein the ORF encodes the amino acid sequence of SEQ ID NO:

90.

4. The composition according to claim 1, further comprising all or a portion of mRNA containing orf1a and / or orf1b, wherein the mRNA encodes SARS-CoV-2 nonstructural protein polymerase and / or RdRp.

5. The composition according to claim 4, wherein the SARS-CoV-2 nonstructural protein polymerase is encoded by SEQ ID NO: 5, and the RdRp is encoded by SEQ ID NO:

6.

6. The composition according to any one of claims 1-5, wherein the mRNA is a chemically modified mRNA sequence comprising one or more N1-methylpseudouridine and / or pseudouridine residues.

7. The composition according to claim 6, wherein all uridine residues in the mRNA are N1-methylpseudouridine or pseudouridine.

8. The composition according to any one of claims 1-5, wherein the mRNA molecule further comprises 3'-UTR and 5'-UTR.

9. The composition according to any one of claims 1-5, wherein the nanoparticles further comprise lipids.

10. The composition according to claim 9, wherein the lipid is a cationic lipid.

11. A method for preparing the composition of any one of claims 1-8, comprising expressing the SARS-CoV-2 protein in a linear in vitro transcription system.

12. A method for preparing the composition of any one of claims 1-8, comprising expressing the SARS-CoV-2 protein using a plasmid DNA vector delivery system.

13. A method for preparing the composition of claim 1, comprising contacting mRNA containing the ORF with HKP, wherein the mRNA and the HKP self-assemble into nanoparticles.

14. Use of the composition of any one of claims 1-9 in the preparation of a medicament for treating a person suffering from SARS-CoV-2 infection.

15. The application according to claim 14, wherein the drug is formulated to be nebulized by a nebulizer inhalation system before or during administration.

16. The application of claim 15, wherein the nebulizer inhalation system is a portable nebulizer for whole respiratory tract drug delivery.

17. The application according to claim 14, wherein the drug is formulated for administration by subcutaneous injection.

18. The application according to claim 14, wherein the drug is formulated for administration by intramuscular injection.

19. The application according to claim 14, wherein the drug is formulated for administration by intraperitoneal injection.

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