中文摘要：

目的构建柔嫩艾美尔球虫病毒RNA依赖RNA聚合酶（RDRP）蛋白部分区段的原核表达重组质粒,并在大肠埃希菌中表达目的蛋白。方法根据柔嫩艾美尔球虫病毒RDRP基因序列设计引物,以病毒基因组dsRNA为模板,RT-PCR扩增目的片段;扩增产物与pMD-18T克隆载体相连接,经测序鉴定序列正确后,将该质粒酶切,酶切目的片段与原核表达载体pET-28a连接构建原核表达质粒,转化入Transetta（DE3）中,经IPTG诱导表达重组蛋白,用SDSPAGE和Western blot分析目的蛋白表达。结果成功克隆柔嫩艾美尔球虫病毒RDRP基因部分序列,其长度为1 330bp,DNA测序表明重组原核表达质粒构建成功,推导的编码氨基酸序列与GenBank中布氏艾美尔球虫病毒RDRP氨基酸序列同源区段同源性为36%。经SDS-PAGE和Western blot分析,柔嫩艾美尔球虫病毒部分RDRP蛋白得到表达,表达蛋白的分子质量单位为52ku,与理论值相符,且该重组蛋白可被His标签单克隆抗体识别。结论构建的重组原核表达载体pET-28a-RDRP在大肠埃希菌中高效表达柔嫩艾美尔球虫病毒RDRP蛋白部分区段,为进一步研究该病毒奠定了基础。

英文摘要：

Objectives To construct a recombinant plasmid and to express part of the Eimeria tenella virus RDRP protein in Escherichia coli. Methods A pair of primers was designed and synthesized in accordance with the nucleotide sequence of E.tenella virus RDRP.A 1330-bp gene fragment of virus genomic dsRNA was amplified via RT-PCR and cloned into the vector pMD18-T.Identified by sequencing,the gene fragment was subcloned into the recombinant plasmid PET-28a-RDRP.The recombinant plasmid was then transferred into E.coli Transetta（DE3）for expression via induction with IPTG.The expressed product was identified using SDS-PAGE and Western blotting. Results A portion of the E.tenellavirus RDRP gene was successfully amplified with a length of 1 330bp,and the predicted amino acid sequence of E.tenellavirus RDRP had a similarity of 36%to that of the E.brunetti RNA virus.The recombinant E.tenella RDRP protein was highly expressed and had a molecular weight of 52ku according to SDS-PAGE and Western blotting. Conclusion Results indicated a portion of the E.tenellavirus RDRP protein was successfully expressed in Transetta（DE3）,laying the foundation for further study of the E.tenellavirus.