畜牧兽医学报 ›› 2005, Vol. 36 ›› Issue (9): 964-968.doi:

• 研究简报 • 上一篇    下一篇

猪A组轮状病毒Vp7基因与乳酸菌非抗性表达载体的重组及在大肠杆菌中的表达

王春凤;王会岩;于守平;刘尚高   

  1. 1. 吉林农业大学动物科学与技术学院,长春 130118;2.中国农业大学动物医学院,北京 100094
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2005-09-25 发布日期:2005-09-25

Expression of Recombinant Vp7 Gene of Porcine Rotavirus A with Non-antibiotic Lactobacillus Vector in Escherichia coli

WANG Chun-feng; WANG Hui-yan; YU Shou-ping; LIU Shang-gao   

  1. 1.College of Animal Science and Technology, Jilin Agricultural University, Changchun 130118, China;2.College of Veterinary Medicine,China Agricultural University, Beijing 100094, China
  • Received:1900-01-01 Revised:1900-01-01 Online:2005-09-25 Published:2005-09-25

Abstract: The antigenic determinants of Vp7 gene of porcine rotavirus A were amplified from cells infected with rotavirus by the reverse transcription-polymerase chain reaction (RT-PCR), and a 981bp cDNA segment was acquived. Using T-A cloning technique, the PCR product was cloned into pGEM-T vector. Cloning plasmid pGEM-T-Vp7 and the prokaryotic expression shuttle vector between E.coli and Lactobacillus pW425t were digested by SacI and KpnI double enzymes, respectively. The purified Vp7 gene was subcloned into the expression vector pW425t. Thus,the recombinant pW425t-Vp7 was constructed, and then was transformed into the competence thyA gene-mutant E.coli X13. Treated lysates of bacterium were loaded directly onto SDS-PAGE, on which approximately 37.07 ku exogenous protein was observed and it was about 15% of the total protein . The protein was further analyzed using Western blot, which indicated that the protein was reactive with the antibody of rotavirus A. The results lay foundation for further studies on the Lactobacillus subunit vaccine and DNA vaccine of Vp7 gene in prevention of porcine rotavirus.