| 摘要: |
| 针对水疱性口炎病毒(VSV)N基因设计特异性引物，在引物中分别加入EcoRI和XhoI酶切位点。RT-PCR扩增后得到目的基因，将其克隆到表达载体pET30c，连接产物转化于DH5a菌株，在含Kan+ 的平板上37 ℃培养24 h，然后挑取白色菌落，鉴定为阳性者，转化于BL21菌株，再经异丙基硫代-β-D-半乳糖苷(IPTG)诱导获得高效表达。表达融合蛋白分子质量为53 ku(目的蛋白大约47 ku)，同预期蛋白大小相近。Western blot检测表明，其能与本病毒阳性血清发生特异性反应，表达蛋白有望成为有价值的VSV诊断抗原。 |
| 关键词: 水疱性口炎病毒 N基因 原核表达载体 |
| DOI： |
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| 基金项目:农业部外来病研究项目(动物疫情监测与防治经费科目编码:070104) |
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| Construction and expression of vesicular stomatitis virus N gene in prokaryotic vector |
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| Abstract: |
| The N gene of VSV was amplified by RT-PCR.After the amplified fragment was cloned into the expression vector pET30c,the insert position,the size and the reading frame of the insertion were identified by PCR,restriction digestion and sequence analysis of the recombinant plasmids.SDS-PAGE and Western blot showed the recombinant plasmids induced by IPTG could express the N gene of VSV,the expression protein could be recognized by the positive serum of VSV.The recombinant N protein is potentially valuable antigen for serological test and immunoprophlaxis of VSV. |
| Key words: vesicular stomatitis virus N gene prokaryotic expression vector |
