猪附红细胞体SRP54基因的克隆及真核表达质粒的构建

Cloning and construction of eukaryotic expression plasmid of the gene encoding the 54 ku subunit of signal recognition particle from Eperythrozoon suis

为了研究猪附红细胞体信号识别颗粒54(SRP54)基因的功能,试验根据GenBank上发表的猪附红细胞体KI_3806全基因组序列(登录号为NC_015153.1)中信号识别颗粒(SRP)蛋白基因设计合成1对特异性引物,采用PCR方法扩增猪附红细胞体SRP54基因片段,将扩增产物克隆至pMD18-T载体上,重组质粒经PCR和酶切鉴定后测序,并将SRP54基因与pVAXⅠ真核表达载体连接。结果表明:克隆的SRP54基因片段长1 173 bp,与GenBank中原序列同源性为97%,构建的真核表达质粒pVAXⅠ-SRP54经PCR和酶切鉴定正确,为猪附红细胞体SRP54基因的后续研究奠定了基础。

To study the function of the gene encoding the 54 ku subunit of signal recognition particle （ SRP54 ） of Eperythrozoon suis, a pair of specific primers were designed and synthetized based on the complete genome sequence of Eperythrozoon suis KI_3806 published in GenBank （ NC_015153.1 ）. The SRP54 gene fragment from Eperythrozoon suis was amplified by PCR and cloned to the pMD18 - T simple vector. The re- combinant plasmid was identified by PCR and restriction enzyme digestion, which was sequenced. Then the SRP54 gene was connected with eu karyotic expression vector pVAX I. The results showed that the gene fragment of SRP54 was 1 173 bp, displayed 97% homology with corre sponding sequences in GenBank. The eukaryotic expression plasmid of pVAX I SRP54 was constructed correctly to be identified by PCR and restriction enzyme digestion, which was laid the foundation for the follow up studies of the SRP54 gene of Eperythrozoon suis.