China Animal Husbandry and Veterinary Medicine ›› 2025, Vol. 52 ›› Issue (4): 1784-1795.doi: 10.16431/j.cnki.1671-7236.2025.04.030

• Preventive Veterinary Medicine • Previous Articles     Next Articles

Cloning and Bioinformatics Analysis of VP7 Gene of the Genotype G8 Bovine Rotavirus

MIAO Shukui1,2, MI Xiaoyun1,2, WEI Jie1,2, WEI Yurong1,2, HAILIQIEMU·Maimaitiyiming1,2   

  1. 1. Institute of Veterinary Medicine, Xinjiang Academy of Animal Sciences (Animal Clinical Medicine Research Center, Xinjiang Academy of Animal Sciences), Urumqi 830013, China;
    2. Xinjiang Key Laboratory of Animal Infectious Diseases, Urumqi 830013, China
  • Received:2024-07-31 Published:2025-03-29

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Abstract: 【Objective】 The purpose of this study was to clone VP7 gene of genetype G8 Bovine rotavirus (BRV) isolated in Xinjiang,and analyze the molecular genetic characteristics of the VP7 protein,so as to provide a theoretical basis for studying its biological functions and new vaccines.【Method】 The viral genome RNA was extracted from the 6th passages of BRV isolate in Xinjiang,and VP7 gene fragment of BRV was amplified by RT-PCR,cloned into pMD19-T vector,and then transformed into Escherichia coli DH5α competent cells.The plasmid was extracted,digested and sequenced.VP7 protein was analyzed using bioinformatics software,including physicochemical properties,transmembrane structure,hydrophilicity/hydrophobicity analysis,subcellular localization,signal peptides,glycosylation sites,phosphorylation sites,secondary structure,tertiary structure homology modeling,and prediction of B and T cell antigen epitopes.【Result】 VP7 gene nucleotide sequence was successfully obtained by RT-PCR amplification, with a total length of 1 062 bp, and then submitted to NCBI to obtain GenBank accession No.: OR136878.1. This sequence had the highest identity (89.4%) with the nucleotide sequence of RVA/Cow-wt/TUR/Amasya-1/2015/G8P[5] strain (GenBank accession No.:KX212865.1),the two strains belonged to genotype G8 of serotype A.Bioinformatics analysis suggested that the BRV VP7 protein was a hydrophobic unstable protein with two transmembrane regions and without signal peptide.VP7 protein was located in endoplasmic reticulum membrane of the host cell.It contained 2 N-glycosylation sites and 132 phosphorylation sites.The secondary structure of VP7 protein was mainly consisted of alpha helix and random coil,which accounted for 36.20% and 35.28%,respectively.VP7 protein could be homologously modeled with the template (SMTL ID:8bp8.1) in the SWISS-MODEL database,with a sequence identity of 82.82%,which indicated a high coincidence rate between them.The GMQE,QMEAN and Ramachandran favored values of the model were 0.75,0.79 and 95.52%,respectively,indicating that the spatial conformation was reasonable and the model was accurate and reliable.The VP7 protein possessed several B and T cell antigenic epitopes.While the epitope length was 16 amino acids and the score exceeds 0.80,there were 12 B cell epitopes.While the epitope length was 9 amino acids,with 4 alleles selected and a score greater than 0.5,there were 6 cytotoxic T lymphocyte (CTL) epitopes.While the epitope length ranged from 12 to 18 amino acids,with 4 alleles selected and a score greater than 0.8,there were 11 helper T lymphocyte (HTL) epitopes.【Conclusion】 The complete nucleotide sequence of VP7 gene of genetype G8 BRV isolated in Xinjiang was successfully obtained for the first time.VP7 protein was an unstable protein with two transmembrane regions and without signal peptide. It was located in endoplasmic reticulum membrane of the host cell,and possessed several B and T cell antigenic epitopes.The experimental results laid the foundation for the epidemic,diagnosis,virus-host interaction,and the research of genetically engineering vaccine targeting VP7 protein of BRV.

Key words: Bovine rotavirus (BRV); VP7 gene; cloning; bioinformatics

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