新疆加工番茄上南方番茄病毒外壳蛋白基因的克隆与原核表达

Cloning and prokaryotic expression of coat protein gene of Southern tomato virus from processing tomato in Xinjiang

【目的】克隆南方番茄病毒(STV)的外壳蛋白(CP)基因,构建其原核表达载体并进行诱导表达,为制备检测该病毒的高效价血清提供参考。【方法】利用一步法RT-PCR从新疆加工番茄上克隆STVCP基因,将其连接到原核表达载体pET-28a(+)上,获得重组质粒pET-28a-STV CP。将重组质粒转化大肠杆菌BL21后用IPTG进行诱导表达。【结果】成功克隆了STVCP基因,其长度为1 134bp。构建了原核重组表达质粒pET-28a-STV CP,其在1mmol/L IPTG诱导下,成功表达出分子质量约47ku的蛋白。【结论】成功克隆了STV CP基因,并诱导了pET-28a-STV CP重组蛋白的原核表达。

【Objective】This paper aimed to clone the coat protein(CP)gene of Southern tomato virus(STV),and construct and express its prokaryotic expression vector,providing reference for preparation and detection of high titer serum of the virus.【Method】The STV CPgene was cloned by One-Step RT-PCR and connected with pET-28a(+)to obtain pET-28a-STV CP.Then the recombinant plasmid was transformed into E.coli BL21 and induced by IPTG.【Result】The STV CPgene was cloned successfully with the size of 1 134 bp and prokaryotic expression vector pET-28a-STV CP was also constructed.The CPprotein with molecular weight of 47 ku was highly expressed after being induced by 1mmol/L IPTG.【Conclusion】STVCPgene was successfully cloned,and the expression of recombinant protein was induced successfully.