[目的]筛选出目的蛋白能够高效表达的重组质粒.[方法]利用原核表达载体pET-32a(+)和pET-28a(+)成功构建了番茄黄化曲叶病毒(Tomato Yellow Leaf Curl Virus,TYLCV)外壳蛋白(Coat Protein,CP)基因的重组质粒p32a-CP和p28a-CP,并通过PCR和双酶切鉴定了序列连接的正确性；再分别将2个载体转化至BL21(DE3)中,采用不同浓度的IPTG对其进行诱导表达,并进行SDS-PAGE电泳检测.[结果]测序结果显示TYLCV-CP基因已定向插入p32a-CP和p28a-CP中；SDS-PAGE电泳结果显示分子量约为50 kD的目的蛋白在重组载体p32a-CP中得到了高效表达,而在重组载体p28a-CP中未表达.[结论]为下一步的抗体制备及TYLCV的免疫学检测奠定了基础.%[Objective] The aim was to screen out recombinant plasmids, in which target protein could express effectively. [Method] The re-combinant plasmid p32a-CP and p28a-CP of coat protein (CP) of tomato yellow leaf curl virus (TYLCV) were constructed successfully by u-sing prokaryotic expression vector pET32a ( + ) and pET-28a ( + ) , and then the correctness of linker sequence was identified by PCR and double digestion. The two constructed vectors were transformed into E. Coli BL21 (DE3) , induced by different concentrations of IPTG and detected by SDS-PAGE. [ Result] PCR and double digestion results showed that TYLCV-CP was correctly inserted into the recombinant plasmid p32a-CP and p28a-CP. After these plasmid were transformed into E. Coli BL21 (DE3) , the target protein about 50 kD in size was expressed highly in p32a-CP while it was not obtained in p28a-CP under different concentration of IPTG by SDS-PAGE. [Conclusion] The research result laid a foundation for antibody preparation and immunological detection of TYLCV.
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