CN113061581A - Goat-derived bovine herpes virus type 1 strain and application thereof - Google Patents

Abstract

The invention belongs to the technical field of animal infectious disease prevention and treatment, and particularly relates to a Goat-derived bovine herpes virus type 1 strain BoHV-1/Goat/CHN/HB 01/2016. The virus strain is separated from excrement of diarrhea goats, and the replication capacity of the virus strain is weakened through virus titer, one-step growth curve, plaque size and other results in vitro MDBK cells. The genome of the virus has the full length of 133,971bp, the GC content is up to 71 percent, and the virus belongs to a BoHV-1.1 subtype through comparison analysis of a gC fragment, wherein specific mutations exist in gB, gK, gM and VP8 proteins and can be related to the pathogenicity of strains. After the strain attacks the cattle, the virulence of the strain is weakened through clinical symptom scoring, nasal cavity detoxification and lung tissue pathological changes. The invention lays an important foundation for researching the pathogenicity and cross-species transmission mechanism of BoHV-1, and provides an important material for clinical vaccine development.

Description

Goat-derived bovine herpes virus type 1 strain and application thereof

Technical Field

The invention belongs to the technical field of animal infectious disease prevention and treatment, and particularly relates to a goat-derived bovine herpes virus type 1 strain.

Background

Bovine herpes virus type 1 (Bovine herpesvirus-1, BoHV-1) belongs to the genus varicella (Varicelovirus) of the family Herpesviridae (Herpesviridae) sub-family α -herpesvirus (Alphaherpesviridae), also known as Bovine infectious rhinotracheitis virus (IBRV). BoHV-1 is the etiological agent of infectious bovine rhinotracheitis, and can cause cattle to develop acute, febrile and contact infectious diseases, and the main clinical manifestations include upper respiratory tract and tracheitis, mucositis, hyperpyrexia, nasal fluid, dyspnea, abortion, etc. After BoHV-1 infection, the body can be immunosuppressed, secondary bacterial or other viral infection can be caused, and severe pneumonia is caused, which is called Bovine respiratory disease syndrome (BRDC), also called as heat of transport, and causes great loss to the cattle industry in the world.

BoHV-1 primarily infects cattle and buffalos, and serological evidence also suggests infection of other artiodactyls. The positive rate of neutralizing antibodies of BoHV-1 in Turkey goat flock was 20.9%; the positive rate of the neutralizing antibody in the sheep is 10.7-28.4%; the seropositivity rates of Egypt sheep and goat BoHV-1 infection are 23.8% and 27.6%, respectively; in Canada, the neutralization antibody positive rate of sheep BoHV-1 is 10.8%; BoHV-1 seropositivity in goats was determined to be 13.7% in the Korean neutralization test. The research shows that: there is a significant correlation between infection between goats and cattle compared to sheep, and goats can therefore be considered as potential storage hosts for BoHV-1. The diversity of the viral host adds difficulty to the control and prevention of the disease, and few studies on the isolation and identification of goat-derived BoHV-1 strains currently limit the mechanism of cross-species transmission of BoHV-1 between goats and cattle. The inventor obtains a goat source BoHV-1 from diarrhea goat feces through separation and identification of virus for the first time, and verifies that the virus has natural attenuated virulence in vivo and in vitro, so that a good natural material is provided for researching the pathogenic and cross-species transmission mechanism of BoHV-1 and developing vaccines.

Disclosure of Invention

The invention aims to provide a goat-derived Bovine herpes virus-1 (Bovine herpesvirus-1) strain, which can be used as a target for researching BoHV-1 pathogenesis and cross-species transmission mechanism, can also be used as a better material for vaccine development, and has important significance for the prevention and control of Bovine herpes virus.

The technical scheme of the invention is as follows:

the applicant obtains a BoHV-1 strain by separating, identifying and purifying the feces of diarrhea goats from a certain Goat farm of Huanggang, Hubei province in 5 months in 2016, the applicant names the strain as bovine herpes virus 1 BoHV-1/coat/CHN/HB 01/2016, and the strain is preserved in China Center for Type Culture Collection (CCTCC) of Wuhan university, Wuhan city, Hubei province in 12 months and 11 days in 2020, with the preservation number of CCTCC NO: v202092.

In vitro MDBK cells, the replication capacity of the strain is proved to be lower than that of a reference strain BoHV-1 Cooper through the results of virus plaques, one-step growth curves, virus titer and the like.

Extracting the whole genome of the virus to perform high-throughput sequencing to obtain a complete virus genome sequence with the genome total length of 133,971bp and the GC content of 71 percent. The virus belongs to the BoHV-1.1 subtype by alignment analysis of the gC fragment. In comparison with the reference strain Cooper, there are specific mutations in the gB, gK, gM, VP8 proteins that may be correlated with the virulence of the strain.

After the cattle is attacked by the virus, the pathogenicity of the strain is lower than that of a reference strain Cooper through clinical symptom scoring, nasal cavity detoxification and lung tissue pathological change.

The invention has the following advantages:

the bovine herpes virus type 1 strain is separated and identified from a diarrhea goat excrement sample for the first time, and the strain shows attenuated virulence, so that an important basis is laid for researching the virulence and cross-species propagation mechanism of the BoHV-1 virus, and an important material is provided for developing a bovine infectious rhinotracheitis vaccine with better clinical safety.

The detailed technical scheme is described in the detailed description.

Drawings

FIG. 1: is a PCR gel map for identifying gG and TK genes of BoHV-1 in a goat diarrhea sample. Description of reference numerals: lanes 1-3 detect the TK gene, and lanes 4-6 detect the gG gene. Lanes 1 and 4 are goat diarrhea samples, lanes 2 and 5 are reference strain Cooper strains as positive controls, lanes 3 and 6 are negative controls, and lane 7 is DL 2000DNA marker.

FIG. 2: is a cytopathic map formed by the BoHV-1 HB01 strain with a scale of 400 μm and with reference numbers indicating: fig. 2A is a cytopathic map formed by goat-derived HB01 strain, and fig. 2B is a negative control map.

FIG. 3: is a morphological map of BoHV-1 HB01 virus particles observed under an electron microscope.

FIG. 4: is the titer stability of the BoHV-1 HB01 virus during passaging.

FIG. 5: is a BoHV-1 HB01 virus growth graph, infecting the virus at 0.1 MOI.

FIG. 6: is a comparison of the size of plaques formed by the BoHV-1 HB01 virus.

FIG. 7: is a gB gene evolutionary tree of BoHV-1 HB01 virus.

FIG. 8: is the gC gene evolutionary tree of the BoHV-1 HB01 virus.

FIG. 9: the temperature change of boHV-1 HB01 virus attacking cattle after the virus attack.

FIG. 10: is the change of clinical symptom scores after BoHV-1 HB01 virus challenge cattle.

FIG. 11: is the daily gain of BoHV-1 HB01 virus after challenge of cattle.

FIG. 12: is to detect the nasal detoxification after BoHV-1 HB01 virus challenge cattle.

FIG. 13: is the histopathological result of the lung of the cattle attacked by the BoHV-1 HB01 virus. Description of reference numerals: FIG. 13A is the pathological observation result of the lung of a bovine after the challenge with BoHV-1 HB01 strain, FIG. 13B is the pathological observation result of the lung of a bovine after the challenge with BoHV-1 Cooper strain, and C in FIG. 13 is the pathological observation result of the lung of a bovine inoculated with DMEM, and the scale in the graph is 50 μm.

Detailed Description

The present invention will be described in detail below with reference to specific examples.

Example 1: isolation and identification of strains

1. Isolation of strains

Collecting a stool sample of a diarrhea goat in a certain goat farm in Huanggang city, preserving at low temperature and rapidly transporting to a laboratory. Samples are centrifuged at 8000R/min for 10min, 200 mu L of supernatant is taken, virus DNA in the samples is extracted by using a DNA extraction kit (purchased from Beijing Quanyu Co., Ltd.), and two pairs of primers gG-F/R, TK-F/R are designed according to the BoHV-1 strain sequence (genome GenBank accession number is AJ004801) for PCR identification. Primers were synthesized by commercial organisms, and the sequences of the primers are shown in Table 1. The resulting target band was as expected (FIG. 1), and BoHV-1 was identified as positive in the goat diarrhea sample.

Table 1: primers for detecting gG and TK genes of BoHV-1 in clinical samples

To a 6-well plate full of MDBK cells, 400. mu.L of the supernatant of the disease material for BoHV-1 positive identification by PCR as described above was added, and the mixture was incubated at 37 ℃ for 60 min. Discarding supernatant, rinsing with sterile PBS for 3 times, adding 2mL DMEM medium containing 2% fetal calf serum, and standing at 37 deg.C with 5% CO₂Cultured in an incubator. After inoculation of the disease material for 18-24h, obvious cytopathy can be observed, the cells become round and have high refractivity, and the cells are gathered into a grape-shaped cluster (figure 2A), and negative control cells have no obvious change (figure 2B).

2. Identification of strains

The virus culture supernatant is subjected to PCR identification of gG and TK fragments after DNA is extracted according to the method, and an expected obvious band is obtained. The PCR product was recovered and purified using agarose gel DNA recovery kit (purchased from Tiangen Biotechnology Ltd.), then ligated to TOPO vector (purchased from Beijing Ederlie Ltd.), the ligation product was transformed into DH 5. alpha. E.coli competent plasmid, inoculated to ampicillin resistant common plate, cultured overnight at 37 ℃, selected for suitable single colony, PCR-identified and the positive single colony was sent to commercial organism for sequencing. The obtained gG (SEQ ID No.1) and TK (SEQ ID No.2) fragments were aligned in Basic Local Alignment Search Tool (BLAST) in GenBank, and both gG and TK were found to have 100% similarity to the corresponding sequences of BoHV-1.

After a large amount of virus was propagated, 8000 Xg of the virus solution was centrifuged for 30min, and the supernatant was filtered with a 0.22 μm filter to remove cell debris, followed by ultracentrifugation of the virus solution at 4 ℃ at 25,000 Xg for 2 h. Discarding the supernatant, fixing the precipitate with glutaraldehyde, dropping the sample on a copper net for adsorption for 5min, dropping 20g/L phosphotungstic acid after drying for negative dyeing for 1min, and then taking a picture under a Hitachi H-7000FA electron microscope for observation. The virus size was approximately 160nm, and it appeared as a spherical, enveloped, and the nucleocapsid of the virion was observed to have a regular hexagonal structure, conforming to the basic characteristics of a BoHV-1 virion (fig. 3).

In conclusion, the isolated virus was identified as bovine herpes virus type 1 and the Applicant named it BoHV-1/Goat/CHN/HB 01/2016.

Example 2: biological Properties of the BoHV-1/Goat/CHN/HB01/2016 Strain

1. Subculture stability of strains

The strain BoHV-1 HB01 was serially passaged through MDBK cells to passage 10, and PFU (plate Forming Unit) was determined for each passage. Diluting the virus to be detected by 10 times, and taking 10^-3，10^-4，10^-5，10^-6And 10^-75 dilutions, plating MDBK cells, 3 wells per dilution, 0.2mL per well, 5% CO at 37 ℃₂Incubate in the incubator, aspirate the virus solution after 2h, and add 1ml of agarose overlay fluid (1% serum in 2 × DMEM: 2% agarose 1:1) per well. After the culture medium is solidified, placingCulturing in 37 deg.C incubator, 48 hr, fixing with 10% neutral formaldehyde for 24 hr, staining cells with crystal violet (0.35%, w/v), washing with water after 20min, oven drying, calculating number of plaques, and calculating virus titer (PFU) according to formula. The virus titer calculation formula is as follows: viral content per ml-average number of plaques triplicated at the same dilution × 5

The result shows that the virus gradually increases the toxic valence to the 4 th generation after MDBK cell passage and is stable at 10%⁷PFU/mL or so (FIG. 4).

2. Growth Curve plotting of strains

Infecting MDBK cells with 0.1MOI virus, harvesting cultures at 0h, 3h, 6h, 9h, 12h, 15h, 18h, 21h, 24h, 30h, 36h, 42h, 48h, 54h, 60h, 66h and 72h after infection respectively to a refrigerator at minus 80 ℃, repeatedly freezing and thawing twice, measuring virus toxicity according to the method, and finally drawing a growth curve. BoHV-1 Cooper strain was used as a reference control strain.

The results show that the overall growth trend of the strain is substantially consistent with that of the reference strain BoHV-1 Cooper. The titer of the BoHV-1 HB01 strain rises faster during 15-36h of infection and reaches a plateau after 42h (10 h)^7.3PFU/mL) (FIG. 5).

3. Plaque size of strains

The plaque diameter of the virus was observed under a microscope after fixed staining according to the titer determination method. 40-50 plaques were selected for measurement calculation, and the plaque diameter formed by the strain was 200 μm on average, and the significance was less than that of the reference strain BoHV-1 Cooper (p <0.0001) (FIG. 6).

Analysis of the biological properties of the virus indicates that the isolated strains of the invention exhibit hypotoxicity in vitro.

Example 3: genomic sequencing of BoHV-1/Goat/CHN/HB01/2016 strain

Extracting the genome of the isolate by using a virus DNA extraction kit (purchased from QAIGEN, Germany), detecting the integrity and concentration of the genome extraction, and sending the qualified DNA sample to Shenzhen Hua Dagen science and technology service GmbH for sequencing and assembling. Homologous gene prediction and annotation was performed based on the reference sequence AJ 004801.

The strain obtained by separation, identification and purification is sequenced and then data assembly is carried out, and finally 1 whole genome sequence is obtained. The result shows that the genome has the full length of 133,971bp, the GC content is up to 71 percent, 73 open reading frames are contained, and 73 proteins are coded. The gB sequences of the isolated and identified BoHV-1 HB01 strain, bovine herpes viruses (BoHV-1 and BoHV-5) and goat herpes virus (CapHV-1) were subjected to evolutionary analysis using MEGA7.0 software, and the analysis was calculated by the orthodox method, giving a Bootstrap value of 1000 times. The results are shown in FIG. 7, and the evolutionary analysis shows that the goat-derived isolate identified by the present invention belongs to bovine herpes virus, and is far away from goat herpes virus CapHV-1. The 451bp sequence of gC was selected for BoHV typing analysis and the results are shown in FIG. 8. The result shows that the strain separated and identified by the invention belongs to BoHV-1.1.

The UL53(SEQ ID No.3), UL27(SEQ ID No.4), UL10(SEQ ID No.5) and UL47(SEQ ID No.6) genes of the strains were translated into glycoprotein gK, gB, gM and VP8 amino acid sequences, respectively, and compared to the published BoHV-1 strain. The results showed that the gK protein (SEQ ID No.7) was missing 6 amino acids (33R, 38L, 202R, 231A, 267A, 274A, respectively) compared to the reference strain Cooper strain. gB (SEQ ID No.8) has a deletion of tyrosine (Y) at position 176, compared to the reference strain Cooper strain. gM (SEQ ID No.9) has the 169-172 continuous mutation (LAPP > AGAA), the 313-317 continuous mutation (RAAAR > GLPLG) and the 406-412 insertion mutation (RPPTIPL > ARQPYR) compared with the reference strain Cooper strain. VP8(SEQ ID No.10) has consecutive mutations at positions 13-15 (RAV > PRR), deletion mutation at positions 33-47 (AGRPARCGRRGRGA > LLDALRAADAEAAER), and positions 365-375 and 381-396 compared to the Cooper strain of the reference strain.

Example 4: virulence of the BoHV-1/Goat/CHN/HB01/2016 strain

The 9 test calves were randomly divided into 3 groups and tested negative for herpes virus type 1. Group 1 inoculation 4X 10^7.0TCID₅₀BoHV-1 HB01, group 2 seed 4X 10^7.0TCID₅₀BoHV-1 Cooper (R), control group was inoculated with 4mL of DMEM in a nasal inoculation. Within 14 days after inoculation, each timeBody temperature was measured 2 times a day, clinical symptoms were recorded, nasal, ocular and anal swabs were collected for virus detection, and serum was collected at 0, 7, 14, 21, 28 for antibody detection. After cessation of detoxification of nasal cavities was determined on day 18 post inoculation, dexamethasone was injected intramuscularly at 2.5mg/kg, followed by continued daily collection of nasal, ocular and anal swabs. On day 28 after inoculation, all experimental cattle were subsequently euthanized and tissue and organ samples were collected. Cattle were weighed before inoculation and before euthanasia.

The body temperature of the cattle with the toxin attacking increased significantly, reaching 40 ℃ at day 4, and then decreased gradually, but still higher than the normal body temperature, and the body temperature of the cattle with HB01 group was significantly lower than that of the Cooper group (FIG. 9). All of the challenged cattle exhibited clinical symptoms (marked nasal and ocular secretions, lassitude, anorexia) which continued until day 14, during which period the Cooper group cattle had a higher clinical symptom score than the HB01 group (fig. 10). The average weight gain of cattle in the control group is 300 g/day, which is obviously higher than that of 2 challenge groups, while the average weight gain of cattle in the HB01 group is higher than that of Cooper group (FIG. 11).

Viruses were detected in the nasal cavities of cattle in Cooper group and HB01 group (FIG. 12), and the peak period of detoxification was 2-5 days, which reached 10^-4.67TCID₅₀mL and 10^-3.8TCID₅₀mL, until no virus was detected on day 9, and the nasal cavity detoxification of cattle in Cooper group was higher than that in HB01 group as a whole. On day 3, the bull eye swabs of Cooper group and HB01 group all cleared toxins, which respectively reached 10^{- 3.76}TCID₅₀mL and 10^-2.39TCID₅₀mL (Table 2). On day 4, the cow anus swab in Cooper group was detoxified to 10^-2.67TCID₅₀mL, whereas the HB01 group bovine anus swab detoxified for 3 days at 10^-(1.57-1.8)TCID₅₀mL (Table 2).

Table 2: BoHV-1 cow receiving posterior eye swab and anus swab detoxification method

After dexamethasone injection, cattle in the Cooper group developed mild symptoms including nasal and ocular secretions, cough, mental depressionDepression, etc. the nasal cavity toxin expelling time is 3-4 days after injection, and the maximum toxin expelling amount is 10^-1.77TCID₅₀Whereas neither HB01 nor control cattle detected detoxification (FIG. 12).

After killing the cattle on day 28, the lung tissue was fixed with formaldehyde for histopathological observation. The results showed that the alveolar structure of HB01 group was destroyed, the alveolar wall was slightly thickened and a small number of lymphoid cells were present in the alveolar space (FIG. 13A), the normal alveolar structure of Cooper group cattle had disappeared, and a large number of necrotic cells and inflammatory cells were present in the alveolar space (FIG. 13B). Whereas the alveolar structure of the normal control group remained intact and the alveolar space was free of lymphoid cells (FIG. 13C).

In conclusion, the bovine herpes virus 1BoHV-1/Goat/CHN/HB01/2016 strain obtained by the invention has weaker virulence in vitro and in vivo than that of a reference strain BoHV-1 Cooper strain, which lays an important foundation for researching the virulence and cross-species transmission mechanism of the bovine herpes virus type 1, provides an important material for the development of a vaccine with better clinical safety, and has important significance for the prevention and control of the bovine herpes virus type 1.

Appendix: the term in the specification states:

bovine herpes virus type 1BoHV-1/Goat/CHN/HB01/2016 strain, represented by BoHV-1 HB 01.

A reference strain of bovine herpes virus type 1, BoHV-1 Cooper strain (KU198480), is designated BoHV-1 Cooper.

Description of sequence listing:

SEQ ID NO.1 is the gG gene nucleotide sequence of the BoHV-1 HB01 strain, and the sequence length is 1860 bp.

SEQ ID NO.2 is the TK gene nucleotide sequence of BoHV-1 HB01 strain, and the sequence length is 868 bp.

SEQ ID NO.3 is the UL53 gene nucleotide sequence of the BoHV-1 HB01 strain, and the sequence length is 999 bp.

SEQ ID NO.4 is the UL27 gene nucleotide sequence of BoHV-1 HB01 strain, and the sequence length is 2799 bp.

SEQ ID NO.5 is the UL10 gene nucleotide sequence of the BoHV-1 HB01 strain, and the sequence length is 1237 bp.

SEQ ID NO.6 is the UL47 gene nucleotide sequence of the BoHV-1 HB01 strain, and the sequence length is 2219 bp.

SEQ ID NO.7 is the gK protein amino acid sequence of the BoHV-1 HB01 strain, encoding 332 amino acids.

SEQ ID NO.8 is the gB protein amino acid sequence of the BoHV-1 HB01 strain, encoding 932 amino acids.

SEQ ID NO.9 is the gM protein amino acid sequence of the BoHV-1 HB01 strain, encoding 412 amino acids.

SEQ ID NO.10 is the VP8 protein amino acid sequence of BoHV-1 HB01 strain, encoding 738 amino acids.

Sequence listing

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gtgaccacga cctgggcggg cagcacgtac gcggccatta caaaccagta cacggaccgc 600

gtgcccgtgg gcatgggcga gatcacggac ctggtggaca agaagtggcg ctgcctttcg 660

aaagccgagt acctgcgcag cgggcgcaag gtggtggcct ttgaccgcga cgacgacccc 720

tgggaggcgc cgctgaagcc tgcgcggctg agcgcgcccg gggtgcgggg ctggcacacg 780

acggacgatg tgtacacggc gctgggctcg gcggggctct accgcacggg cacctctgtg 840

aactgcatcg tggaagaagt ggaggcgcgc tcggtgtacc cgtacgactc gttcgcgctc 900

tcgaccgggg acattatcta catgtcgccc ttttacgggc tgcgcgaggg cgcgcaccgc 960

gagcacacca gctactcgcc ggagcgcttc cagcagatcg agggctacta caagcgcgac 1020

atggccacgg gccggcgcct caaggagccg gtctcgcgga actttttgcg tacacagcac 1080

gtgacggtag cctgggactg ggtgcccaag cgcaaaaacg tgtgctcgct ggccaagtgg 1140

cgcgaggcgg acgaaatgct gcgagacgag agccgcggga acttccgctt cacggcccgc 1200

tcgctctcgg cgacctttgt gagcgacagc cacaccttcg cgttgcagaa tgtgccgctg 1260

agcgactgcg tgatcgaaga ggccgaggcc gcggtcgagc gcgtctaccg cgagcgctac 1320

aacggcacgc acgtgctgtc gggcagcttg gagacgtacc tggcgcgcgg cggctttgtc 1380

gtggccttcc ggccgatgct cagcaacgag ctggccaagc tgtacctgca ggagctggcg 1440

cgctcgaacg gcacgctcga ggggctgttc gccgccgcgg cgcccaagcc gggcccgcgg 1500

cgcgcgcgcc gcgccgcgcc gtctgcgccc ggcggcccgg gcgcggccaa cgggcccgcc 1560

ggcgacggcg acgccggcgg gcgggtgact accgtgagct cggccgagtt tccggcgctg 1620

cagttcacct acgaccacat ccaggaccac gtgaacacca tgttcagccg cctggccacg 1680

tcctggtgcc tgctgcagaa caaggagcgc gccctgtggg ccgaggcggc taagctcaac 1740

cccagcgcgg cggccagcgc tgcgctggac cgccgcgccg ccgcgcgcat gttgggggac 1800

gccatggccg tgacgtactg ccacgagctg ggcgaggggc gcgtgttcat cgagaactcg 1860

atgcgcgcgc ccggcggcgt ttgctacagc cgcccgccgg tctcctttgc cttcggcaac 1920

gagagcgagc cggtggaggg ccagctcggc gaggacaacg agctgctgcc gggccgcgag 1980

ctcgtggagc cctgcaccgc caaccacaag cgctacttcc gctttggcgc ggactacgtg 2040

tactacgaga actacgcgta cgtgcggcgg gtcccgctcg cggagctgga ggtgatcagc 2100

acctttgtgg acctaaacct cacggttctg gaggaccgcg agttcttgcc gctagaagtg 2160

tacacgcgcg ccgagctcgc cgacacgggt ctgctcgact acagcgagat acagcgccgc 2220

aaccagctgc acgagctccg gttctacgac attgaccgcg tggtcaagac ggacggcaat 2280

atggccatca tgcgagggct cgccaacttc tttcagggcc tgggcgccgt cgggcaggcg 2340

gtgggcacgg tggtgctggg cgccgcgggt gccgcgctct cgaccgtgtc gggcatcgcc 2400

tcgtttattg cgaacccgtt cggcgcgctg gccacggggc tgctggtgct cgccgggctg 2460

gtggccgctt tcctggcgta ccggtacatt tcccgcctcc gcagcaaccc catgaaggcg 2520

ctgtacccga tcaccacgcg cgcgctcaag gacgacgccc ggggcgcaac cgccccgggc 2580

gaggaagagg aggagtttga cgcggccaaa ctggagcagg cccgcgagat gatcaagtat 2640

atgtcgctcg tgtcagcggt cgagcggcaa gagcacaagg cgaaaaagag caacaagggc 2700

ggcccgctgc tggcgacccg gctgacgcag ctcgcgcttc ggcggcgagc gccgccggag 2760

taccagcagc ttccgatggc cgacgtcggg ggggcatga 2799

<210> 5

<211> 1237

<212> DNA

<213> Bovine herpes virus type 1 (Bovine herpesvirus 1)

<400> 5

ttagcggtat ggttggcggg cggggccggg cgtccgcctc gtacacggcc tcctcgtcgt 60

cagaggcgac gccgcggtcc agcagcgccg ctgcgtcgcc ttcgagcggg gccatgccgc 120

ggctgctgcg cgctcgggcg atgtagcgcg ccgcttgctg tcgcgcggcg ctcacgtggc 180

cgtagaaggc gctgcctttg cgccggtggt gcaggcaggc gcgcaccaac cgcagcacga 240

gcatggccac ggccagcagg gcgacgccgg ccagcgccag ctttacccta gcggcagccc 300

ggggggcctg cgtgcgcaca accccccgca gctggtggaa gtagtcgtgc gaggcgaccg 360

cgatgcagct agctgccacc agcgttccaa aggccggccc gggcagcact tgcgtgtagc 420

gcgacaggac tagctcggtc agtaccaaaa agagcacggc ggcgaggcag aagaccgtca 480

cgccggcgac gaccgtttgt gccagcgaca gccccagccc ggcccccagc agctggccga 540

gcatcagggg cgcggtgcca atgatcagcg ccagcaggcc gccggccagg ttgatcatca 600

cggcgcgccc ggggcccagc aggcggtggg cgcgcgggct cctctcgcgg agggcgtgca 660

cggcctgcgc gtactcggcg ccgggcagcc cggcggtgca aaagagcccg acgaaggcgg 720

ccggcggcgc caggtgcgcg aggtaggtgg ccgcggccaa gaccatgatc ttgtgggaca 780

gcagcagcac ggccgcctgc agcgtccagg ccgccagcgc gcccagcagc agcgcacccg 840

ggggcgtggc aagcgtcgcc agcgcgttca tatgatacgc cgccgcaaaa ccccctgagt 900

tttcgcggcg aatgattata gcggccgcga cgtcaaaggc ggcgacggcg agcagaacta 960

cggccgtgta cgcggagaag gccgctgtgg tcggcgtctc caaaaacagc gccgggtggc 1020

gcgcgccgag ctcccgctgc gcccaggcgc cgccgtcggc cgagaggttc cgcgcattgt 1080

aattaactat ggccgcgtag aagcagggga gccccgcgcc cggcgaagcg gcgccgatca 1140

gggtgatcag cagcagcagc gccagcgcgg cgaagacgcc gacctgagcg agccacaagc 1200

gccagtgcac ggcggcaggc tgcgcggagc ccgccat 1237

<210> 6

<211> 2219

<212> DNA

<213> Bovine herpes virus type 1 (Bovine herpesvirus 1)

<400> 6

atggacgccg ctagggatgg gcggcctgag cgccgccgcg ccgtctccgg aacgtaccgc 60

acgcacccgt tccagcgccc ctctgcccgg cggagcgctg gacgccctgc gcgctgcgga 120

cgccgaggcc gcggagcgcc gcgggtccgg cgcccgcggc ctgacttcca gcggcccccg 180

gacgaggaca ccagtgagga cgagaacgtg tatgattaca tcgacggcga tagcagcgac 240

agcgccgacg actatgatag cgattatttt actgctaacc gcggccccaa tcacggcgcc 300

ggcgatgcta tggacacaga cgcaccaccc gagcgcgccc cggaaggggg tgccccgcaa 360

gactacttga cggcccacct gcgcgccatc gaggccctgc ccgagtcagc gccccaccgg 420

agcctgctgg agcgcacggc ccggaccgtg tatgcgcagc agtttccccc gcgcgatttg 480

agtgcgggct ccagggcgcc ggcacagcgc gcgcggcgga gcctccgcgg cttcccgcgt 540

ggcggcgggg gcggccagga acccgggcca gacgacgaag gcgacgacgc cgcagacctg 600

cgcgaggacc ttgtgccaga tgaggcctac gcgcacctag agcgcgacga gcggctgtcg 660

gaagggcccc cgctcctcaa catggaggcg gccgctgcgg ctgcggggga gaggagcgtg 720

gtggaggagc tgtttacgta cgcccctgcc cagcctcagg tagaggtgcc gctgcccagg 780

attttggagg gccgggtgcg gcccagcgcc ttcttcgcgc agatgccgct ggacgcgctg 840

tgccgcacgc cgcccaacga tcagcgcgtg gtgcgcgagc ggcgcgcttg ggagatggcc 900

ggtacgccgc atgggctcct aataaccacg tggagcacgg tggacccgga attctcgatc 960

ggcggcatgt acgtgggcgc ccctgagggc acccggcccc ggctagtgtg gcggcgcgcg 1020

atgaagcagg ccatggcgct gcagtaccgg ctgggggtgg ggggcctgtg ccgagcagta 1080

gacggcgcag catgccgccc actgaggcgc tgctcttttt ggcgggacgc gctgctgcgc 1140

gagtgcgcaa ctgccatttt ttgtcgcggc cggggcgcgc gggcggcgcc gcgccgcctg 1200

ccgcggtcgg cggttgggct gctggcagcc acgcagttca cgccaccgga cgcgtccccc 1260

cacgcgacgc tctttcgcgg ctcaatgggc tccctgattt actggcacga gctgcgcgtg 1320

atgctgactg cggtgccggc cctgtgcgcg cgctacgcgg gcgccgggct gcagtcggcc 1380

gagctgtacc tgctggcgct agcgcactca gaggcgcccg gctacacggc aaatgagcgc 1440

tacgcgctct cggcgtacct gacgctgttt gtagcgctcg cggagcgggc gctgcgctgg 1500

ctgtatctag cgggcgcgca cctgctcggg ccgcacccca cagcggcggc cttccgcgaa 1560

gtgcgcgcca agatcccgta cgagcggctg ccgctaggca gcgcgacgct gcacgacgcc 1620

gaagtggaga cggtggactc ggccaccttc caggaggccc tggcttttag cgcgctggca 1680

catgtttacg gggaggccta cgtagcggtg cggaccgcga cgacgctgct gatggccgag 1740

tacgcggccc acgctgagcg ccgggacgtg cgcgagatga cagcggcctt cctgggcgtg 1800

gggctgatcg cgcagcggct gatgggcagc ctgaacctgc tgctgaactg cgtagccggc 1860

gcagcggtgt acgggggccg gcgtgtgacg gtgcgcgagg gcacgctcgc gcggtacagc 1920

ctgctagcgg acgcggcact gccgctggtg cgcccggtgt ccctggtgga gttctgggag 1980

gcccgcgacg gcgtcatgcg cgagctgcgg ctgcggcccg tggcgagccc gcccctggcc 2040

ggcaagcggc gggtcatgga gctgtacctc tcgctggaca gcatagaggc gctggtcggc 2100

cgcgagccgc taggttcgcg gccggtgctt gggccgctcg tggacatcgc ggaggcgctg 2160

gcggaccacc cgcacctcgt cacgggcgat gggcggggcc cgcgcctggg cggccgcta 2219

<210> 7

<211> 332

<212> PRT

<213> Bovine herpes virus type 1 (Bovine herpesvirus 1)

<400> 7

Met Leu Leu Gly Gly Arg Thr Val Asn Leu Ala Ala Leu Ala Leu Leu

1 5 10 15

Thr Ala His Leu Ala Leu Ala Leu Trp Ala Ala Leu Ala Ala Arg Cys

20 25 30

Gln Arg Cys Ala Cys Val Arg Ala Thr Ala His Asn Gly Ser Leu Arg

35 40 45

Trp Glu Leu Arg Asp Pro Gly Ala Val Tyr Val Trp Gly Gly Ala Asn

50 55 60

Asn Ala Thr Leu Ala Ala Asp Ala Pro Cys Arg His Ala Val Val Gln

65 70 75 80

His Ile Pro Pro Gly Leu Leu Asp Gly Asp Glu Ala Leu His Gly Arg

85 90 95

Val Arg Ala Val Ala Gly Ala Arg Asp Cys Arg Ala Tyr Leu Trp Ser

100 105 110

Ala Gln Ala Arg Ser Ala Leu Leu Ala Trp Leu Leu Tyr Val Ala Phe

115 120 125

Val Tyr Leu Arg Gln Glu Arg Arg Met Phe Gly Ile Cys Arg Asn Asp

130 135 140

Ala Asp Phe Leu Ser Pro Gly Gly Tyr Thr Leu Asn Tyr Ala Ala Ala

145 150 155 160

Ala Leu Ala Ala Val Val Gly His Gly Pro Tyr Thr Lys Leu Ala Arg

165 170 175

Leu Met Cys Glu Leu Ser Ala Arg Arg Arg Ala Leu Ala Val Asn Phe

180 185 190

Arg Leu Asp Pro Leu Gly Cys Ala Trp Arg Pro Arg Ala Ala Leu Pro

195 200 205

Leu Leu Ala Glu Gly Leu Ala Arg Leu Gly Ala Arg Ile Ala Ala Ala

210 215 220

Ser Ser Val Gly Ile Thr His Pro Cys Ala Ala Ala Tyr Pro Leu Tyr

225 230 235 240

Leu Lys Ile Trp Ala Trp Val His Val Ala Leu Phe Ala Gly Leu Glu

245 250 255

Leu Val Ser Leu Leu Tyr Arg Lys Pro Arg Arg Arg Gly Gly Thr Cys

260 265 270

Ala Gly Asp Gly Gly Asp Gly Gly Glu Ser Gly Ile Arg Lys Val Cys

275 280 285

Val Asn Cys Cys Ser Thr Leu Leu Ala Gly Leu Leu Val Lys Ala Leu

290 295 300

Tyr Leu Ala Ala Ile Val Gly Gly Val Ile Ala Leu Leu His Tyr Glu

305 310 315 320

His Asn Leu Arg Leu Arg Leu Leu Gly Ala Gln Thr

325 330

<210> 8

<211> 932

<212> PRT

<213> Bovine herpes virus type 1 (Bovine herpesvirus 1)

<400> 8

Met Ala Ala Arg Gly Gly Ala Glu Arg Ala Ala Gly Ala Gly Asp Gly

1 5 10 15

Arg Arg Gly Gln Arg Arg His Leu Arg Pro Gly Arg Val Leu Ala Ala

20 25 30

Leu Arg Gly Pro Ala Ala Pro Gly Ala Gly Gly Ala Arg Ala Ala Leu

35 40 45

Ala Ala Ala Leu Leu Trp Ala Thr Trp Ala Leu Leu Leu Ala Ala Pro

50 55 60

Ala Ala Gly Arg Pro Ala Thr Thr Pro Pro Ala Pro Pro Pro Glu Glu

65 70 75 80

Ala Ala Ser Pro Ala Pro Pro Ala Ser Pro Ser Pro Pro Gly Pro Asp

85 90 95

Gly Asp Asp Ala Ala Ser Pro Asp Asn Ser Thr Asp Val Arg Ala Ala

100 105 110

Leu Arg Leu Ala Gln Ala Ala Gly Glu Asn Ser Arg Phe Phe Val Cys

115 120 125

Pro Pro Pro Ser Gly Ala Thr Val Val Arg Leu Ala Pro Ala Arg Pro

130 135 140

Cys Pro Glu Tyr Gly Leu Gly Arg Asn Tyr Thr Glu Gly Ile Gly Val

145 150 155 160

Ile Tyr Lys Glu Asn Ile Ala Pro Tyr Thr Phe Lys Ala Tyr Ile Tyr

165 170 175

Lys Asn Val Ile Val Thr Thr Thr Trp Ala Gly Ser Thr Tyr Ala Ala

180 185 190

Ile Thr Asn Gln Tyr Thr Asp Arg Val Pro Val Gly Met Gly Glu Ile

195 200 205

Thr Asp Leu Val Asp Lys Lys Trp Arg Cys Leu Ser Lys Ala Glu Tyr

210 215 220

Leu Arg Ser Gly Arg Lys Val Val Ala Phe Asp Arg Asp Asp Asp Pro

225 230 235 240

Trp Glu Ala Pro Leu Lys Pro Ala Arg Leu Ser Ala Pro Gly Val Arg

245 250 255

Gly Trp His Thr Thr Asp Asp Val Tyr Thr Ala Leu Gly Ser Ala Gly

260 265 270

Leu Tyr Arg Thr Gly Thr Ser Val Asn Cys Ile Val Glu Glu Val Glu

275 280 285

Ala Arg Ser Val Tyr Pro Tyr Asp Ser Phe Ala Leu Ser Thr Gly Asp

290 295 300

Ile Ile Tyr Met Ser Pro Phe Tyr Gly Leu Arg Glu Gly Ala His Arg

305 310 315 320

Glu His Thr Ser Tyr Ser Pro Glu Arg Phe Gln Gln Ile Glu Gly Tyr

325 330 335

Tyr Lys Arg Asp Met Ala Thr Gly Arg Arg Leu Lys Glu Pro Val Ser

340 345 350

Arg Asn Phe Leu Arg Thr Gln His Val Thr Val Ala Trp Asp Trp Val

355 360 365

Pro Lys Arg Lys Asn Val Cys Ser Leu Ala Lys Trp Arg Glu Ala Asp

370 375 380

Glu Met Leu Arg Asp Glu Ser Arg Gly Asn Phe Arg Phe Thr Ala Arg

385 390 395 400

Ser Leu Ser Ala Thr Phe Val Ser Asp Ser His Thr Phe Ala Leu Gln

405 410 415

Asn Val Pro Leu Ser Asp Cys Val Ile Glu Glu Ala Glu Ala Ala Val

420 425 430

Glu Arg Val Tyr Arg Glu Arg Tyr Asn Gly Thr His Val Leu Ser Gly

435 440 445

Ser Leu Glu Thr Tyr Leu Ala Arg Gly Gly Phe Val Val Ala Phe Arg

450 455 460

Pro Met Leu Ser Asn Glu Leu Ala Lys Leu Tyr Leu Gln Glu Leu Ala

465 470 475 480

Arg Ser Asn Gly Thr Leu Glu Gly Leu Phe Ala Ala Ala Ala Pro Lys

485 490 495

Pro Gly Pro Arg Arg Ala Arg Arg Ala Ala Pro Ser Ala Pro Gly Gly

500 505 510

Pro Gly Ala Ala Asn Gly Pro Ala Gly Asp Gly Asp Ala Gly Gly Arg

515 520 525

Val Thr Thr Val Ser Ser Ala Glu Phe Pro Ala Leu Gln Phe Thr Tyr

530 535 540

Asp His Ile Gln Asp His Val Asn Thr Met Phe Ser Arg Leu Ala Thr

545 550 555 560

Ser Trp Cys Leu Leu Gln Asn Lys Glu Arg Ala Leu Trp Ala Glu Ala

565 570 575

Ala Lys Leu Asn Pro Ser Ala Ala Ala Ser Ala Ala Leu Asp Arg Arg

580 585 590

Ala Ala Ala Arg Met Leu Gly Asp Ala Met Ala Val Thr Tyr Cys His

595 600 605

Glu Leu Gly Glu Gly Arg Val Phe Ile Glu Asn Ser Met Arg Ala Pro

610 615 620

Gly Gly Val Cys Tyr Ser Arg Pro Pro Val Ser Phe Ala Phe Gly Asn

625 630 635 640

Glu Ser Glu Pro Val Glu Gly Gln Leu Gly Glu Asp Asn Glu Leu Leu

645 650 655

Pro Gly Arg Glu Leu Val Glu Pro Cys Thr Ala Asn His Lys Arg Tyr

660 665 670

Phe Arg Phe Gly Ala Asp Tyr Val Tyr Tyr Glu Asn Tyr Ala Tyr Val

675 680 685

Arg Arg Val Pro Leu Ala Glu Leu Glu Val Ile Ser Thr Phe Val Asp

690 695 700

Leu Asn Leu Thr Val Leu Glu Asp Arg Glu Phe Leu Pro Leu Glu Val

705 710 715 720

Tyr Thr Arg Ala Glu Leu Ala Asp Thr Gly Leu Leu Asp Tyr Ser Glu

725 730 735

Ile Gln Arg Arg Asn Gln Leu His Glu Leu Arg Phe Tyr Asp Ile Asp

740 745 750

Arg Val Val Lys Thr Asp Gly Asn Met Ala Ile Met Arg Gly Leu Ala

755 760 765

Asn Phe Phe Gln Gly Leu Gly Ala Val Gly Gln Ala Val Gly Thr Val

770 775 780

Val Leu Gly Ala Ala Gly Ala Ala Leu Ser Thr Val Ser Gly Ile Ala

785 790 795 800

Ser Phe Ile Ala Asn Pro Phe Gly Ala Leu Ala Thr Gly Leu Leu Val

805 810 815

Leu Ala Gly Leu Val Ala Ala Phe Leu Ala Tyr Arg Tyr Ile Ser Arg

820 825 830

Leu Arg Ser Asn Pro Met Lys Ala Leu Tyr Pro Ile Thr Thr Arg Ala

835 840 845

Leu Lys Asp Asp Ala Arg Gly Ala Thr Ala Pro Gly Glu Glu Glu Glu

850 855 860

Glu Phe Asp Ala Ala Lys Leu Glu Gln Ala Arg Glu Met Ile Lys Tyr

865 870 875 880

Met Ser Leu Val Ser Ala Val Glu Arg Gln Glu His Lys Ala Lys Lys

885 890 895

Ser Asn Lys Gly Gly Pro Leu Leu Ala Thr Arg Leu Thr Gln Leu Ala

900 905 910

Leu Arg Arg Arg Ala Pro Pro Glu Tyr Gln Gln Leu Pro Met Ala Asp

915 920 925

Val Gly Gly Ala

930

<210> 9

<211> 412

<212> PRT

<213> Bovine herpes virus type 1 (Bovine herpesvirus 1)

<400> 9

Met Ala Gly Ser Ala Gln Pro Ala Ala Val His Trp Arg Leu Trp Leu

1 5 10 15

Ala Gln Val Gly Val Phe Ala Ala Leu Ala Leu Leu Leu Leu Ile Thr

20 25 30

Leu Ile Gly Ala Ala Ser Pro Gly Ala Gly Leu Pro Cys Phe Tyr Ala

35 40 45

Ala Ile Val Asn Tyr Asn Ala Arg Asn Leu Ser Ala Asp Gly Gly Ala

50 55 60

Trp Ala Gln Arg Glu Leu Gly Ala Arg His Pro Ala Leu Phe Leu Glu

65 70 75 80

Thr Pro Thr Thr Ala Ala Phe Ser Ala Tyr Thr Ala Val Val Leu Leu

85 90 95

Ala Val Ala Ala Phe Asp Val Ala Ala Ala Ile Ile Ile Arg Arg Glu

100 105 110

Asn Ser Gly Gly Phe Ala Ala Ala Tyr His Met Asn Ala Leu Ala Thr

115 120 125

Leu Ala Thr Pro Pro Gly Ala Leu Leu Leu Gly Ala Leu Ala Ala Trp

130 135 140

Thr Leu Gln Ala Ala Val Leu Leu Leu Ser His Lys Ile Met Val Leu

145 150 155 160

Ala Ala Ala Thr Tyr Leu Ala His Leu Ala Pro Pro Ala Ala Phe Val

165 170 175

Gly Leu Phe Cys Thr Ala Gly Leu Pro Gly Ala Glu Tyr Ala Gln Ala

180 185 190

Val His Ala Leu Arg Glu Arg Ser Pro Arg Ala His Arg Leu Leu Gly

195 200 205

Pro Gly Arg Ala Val Met Ile Asn Leu Ala Gly Gly Leu Leu Ala Leu

210 215 220

Ile Ile Gly Thr Ala Pro Leu Met Leu Gly Gln Leu Leu Gly Ala Gly

225 230 235 240

Leu Gly Leu Ser Leu Ala Gln Thr Val Val Ala Gly Val Thr Val Phe

245 250 255

Cys Leu Ala Ala Val Leu Phe Leu Val Leu Thr Glu Leu Val Leu Ser

260 265 270

Arg Tyr Thr Gln Val Leu Pro Gly Pro Ala Phe Gly Thr Leu Val Ala

275 280 285

Ala Ser Cys Ile Ala Val Ala Ser His Asp Tyr Phe His Gln Leu Arg

290 295 300

Gly Val Val Arg Thr Gln Ala Pro Arg Ala Ala Ala Arg Val Lys Leu

305 310 315 320

Ala Leu Ala Gly Val Ala Leu Leu Ala Val Ala Met Leu Val Leu Arg

325 330 335

Leu Val Arg Ala Cys Leu His His Arg Arg Lys Gly Ser Ala Phe Tyr

340 345 350

Gly His Val Ser Ala Ala Arg Gln Gln Ala Ala Arg Tyr Ile Ala Arg

355 360 365

Ala Arg Ser Ser Arg Gly Met Ala Pro Leu Glu Gly Asp Ala Ala Ala

370 375 380

Leu Leu Asp Arg Gly Val Ala Ser Asp Asp Glu Glu Ala Val Tyr Glu

385 390 395 400

Ala Asp Ala Arg Pro Arg Pro Pro Thr Ile Pro Leu

405 410

<210> 10

<211> 739

<212> PRT

<213> Bovine herpes virus type 1 (Bovine herpesvirus 1)

<400> 10

Met Asp Ala Ala Arg Asp Gly Arg Pro Glu Arg Arg Arg Ala Val Ser

1 5 10 15

Gly Thr Tyr Arg Thr His Pro Phe Gln Arg Pro Ser Ala Arg Arg Ser

20 25 30

Ala Gly Arg Pro Ala Arg Cys Gly Arg Arg Gly Arg Gly Ala Pro Arg

35 40 45

Val Arg Arg Pro Arg Pro Asp Phe Gln Arg Pro Pro Asp Glu Asp Thr

50 55 60

Ser Glu Asp Glu Asn Val Tyr Asp Tyr Ile Asp Gly Asp Ser Ser Asp

65 70 75 80

Ser Ala Asp Asp Tyr Asp Ser Asp Tyr Phe Thr Ala Asn Arg Gly Pro

85 90 95

Asn His Gly Ala Gly Asp Ala Met Asp Thr Asp Ala Pro Pro Glu Arg

100 105 110

Ala Pro Glu Gly Gly Ala Pro Gln Asp Tyr Leu Thr Ala His Leu Arg

115 120 125

Ala Ile Glu Ala Leu Pro Glu Ser Ala Pro His Arg Ser Leu Leu Glu

130 135 140

Arg Thr Ala Arg Thr Val Tyr Ala Gln Gln Phe Pro Pro Arg Asp Leu

145 150 155 160

Ser Ala Gly Ser Arg Ala Pro Ala Gln Arg Ala Arg Arg Ser Leu Arg

165 170 175

Gly Phe Pro Arg Gly Gly Gly Gly Gly Gln Glu Pro Gly Pro Asp Asp

180 185 190

Glu Gly Asp Asp Ala Ala Asp Leu Arg Glu Asp Leu Val Pro Asp Glu

195 200 205

Ala Tyr Ala His Leu Glu Arg Asp Glu Arg Leu Ser Glu Gly Pro Pro

210 215 220

Leu Leu Asn Met Glu Ala Ala Ala Ala Ala Ala Gly Glu Arg Ser Val

225 230 235 240

Val Glu Glu Leu Phe Thr Tyr Ala Pro Ala Gln Pro Gln Val Glu Val

245 250 255

Pro Leu Pro Arg Ile Leu Glu Gly Arg Val Arg Pro Ser Ala Phe Phe

260 265 270

Ala Gln Met Pro Leu Asp Ala Leu Cys Arg Thr Pro Pro Asn Asp Gln

275 280 285

Arg Val Val Arg Glu Arg Arg Ala Trp Glu Met Ala Gly Thr Pro His

290 295 300

Gly Leu Leu Ile Thr Thr Trp Ser Thr Val Asp Pro Glu Phe Ser Ile

305 310 315 320

Gly Gly Met Tyr Val Gly Ala Pro Glu Gly Thr Arg Pro Arg Leu Val

325 330 335

Trp Arg Arg Ala Met Lys Gln Ala Met Ala Leu Gln Tyr Arg Leu Gly

340 345 350

Val Gly Gly Leu Cys Arg Ala Val Asp Gly Ala Ala Cys Arg Pro Leu

355 360 365

Arg Arg Cys Ser Phe Trp Arg Asp Ala Leu Leu Arg Glu Cys Ala Thr

370 375 380

Ala Ile Phe Cys Arg Gly Arg Gly Ala Arg Ala Ala Pro Arg Arg Leu

385 390 395 400

Pro Arg Ser Ala Val Gly Leu Leu Ala Ala Thr Gln Phe Thr Pro Pro

405 410 415

Asp Ala Ser Pro His Ala Thr Leu Phe Arg Gly Ser Met Gly Ser Leu

420 425 430

Ile Tyr Trp His Glu Leu Arg Val Met Leu Thr Ala Val Pro Ala Leu

435 440 445

Cys Ala Arg Tyr Ala Gly Ala Gly Leu Gln Ser Ala Glu Leu Tyr Leu

450 455 460

Leu Ala Leu Ala His Ser Glu Ala Pro Gly Tyr Thr Ala Asn Glu Arg

465 470 475 480

Tyr Ala Leu Ser Ala Tyr Leu Thr Leu Phe Val Ala Leu Ala Glu Arg

485 490 495

Ala Leu Arg Trp Leu Tyr Leu Ala Gly Ala His Leu Leu Gly Pro His

500 505 510

Pro Thr Ala Ala Ala Phe Arg Glu Val Arg Ala Lys Ile Pro Tyr Glu

515 520 525

Arg Leu Pro Leu Gly Ser Ala Thr Leu His Asp Ala Glu Val Glu Thr

530 535 540

Val Asp Ser Ala Thr Phe Gln Glu Ala Leu Ala Phe Ser Ala Leu Ala

545 550 555 560

His Val Tyr Gly Glu Ala Tyr Val Ala Val Arg Thr Ala Thr Thr Leu

565 570 575

Leu Met Ala Glu Tyr Ala Ala His Ala Glu Arg Arg Asp Val Arg Glu

580 585 590

Met Thr Ala Ala Phe Leu Gly Val Gly Leu Ile Ala Gln Arg Leu Met

595 600 605

Gly Ser Leu Asn Leu Leu Leu Asn Cys Val Ala Gly Ala Ala Val Tyr

610 615 620

Gly Gly Arg Arg Val Thr Val Arg Glu Gly Thr Leu Ala Arg Tyr Ser

625 630 635 640

Leu Leu Ala Asp Ala Ala Leu Pro Leu Val Arg Pro Val Ser Leu Val

645 650 655

Glu Phe Trp Glu Ala Arg Asp Gly Val Met Arg Glu Leu Arg Leu Arg

660 665 670

Pro Val Ala Ser Pro Pro Leu Ala Gly Lys Arg Arg Val Met Glu Leu

675 680 685

Tyr Leu Ser Leu Asp Ser Ile Glu Ala Leu Val Gly Arg Glu Pro Leu

690 695 700

Gly Ser Arg Pro Val Leu Gly Pro Leu Val Asp Ile Ala Glu Ala Leu

705 710 715 720

Ala Asp His Pro His Leu Val Thr Gly Asp Gly Arg Gly Pro Arg Leu

725 730 735

Gly Gly Arg

Claims (5)

1. A Bovine herpes virus type 1 (BoHV-1) strain is named as BoHV-1/Goat/CHN/HB01/2016 and is deposited in China center for type culture Collection with the preservation number of CCTCC NO: v202092.

2. The bovine herpes virus type 1 strain of claim 1 which is: the strain is separated from diarrhea goat excrement and is a goat-derived bovine herpes virus type 1 strain.

3. The bovine herpes virus type 1 strain of claim 2 which is: the amino acid sequences of gK, gB, gM and VP8 proteins of the strain are respectively shown in SEQ ID NO. 7-10.

4. Use of the strain of claim 1 in the study of the pathogenesis and cross-species transmission mechanism of bovine herpes virus type 1.

5. Use of the strain of claim 1 which is attenuated for its virulence in the preparation of a bovine herpes virus type 1 vaccine.

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