鸡贫血病毒VP1基因的克隆及其在大肠杆菌中的表达

Cloning of VP1 Gene of Chicken Infectious Anemia Virus and Its Expression in E.coli

根据鸡贫血病毒Cux_1株基因组序列设计了一对引物,通过PCR扩增出VP1基因。将扩增出的片段克隆到pGM_T载体上,通过测序分析证实该片段与鸡贫血病毒Cux_1株的VP1基因序列一致。然后将克隆化VP1基因亚克隆到PET_32a(+)载体上并进行原核表达。SDS_PAGE和Western Blot分析表明,一个分子质量为70.4 kDa的重组蛋白获得了成功表达,并且鸡贫血病毒阳性血清能和重组蛋白发生反应。

We designed a pair of oligonucleotide primers based on the genomic sequence of CAV Cux-1 strain, and amplified VP1 gene by PCR. The amplification fragment was cloned into pGM-T easy vector. The result of nucleotide sequence analysis demonstrated that the cloned genomic DNA was identical with that deduced from Cux-1 strain of CAV. Then the cloned genomic DNA was subcloned into prokaryotic expression vector PET-32a（ ＋ ）, and the recombinant protein was expressed in E. coli cell. SDS-PAGE and Western Blot analysis showed that a protein with a molecular weight of 70.4 kDa was successfully expressed and CAV-infected chicken serum reacted with the recombinant protein.