【摘要】 以副结核分枝杆菌C-2株基因组DNA为模板,以hsp65基因特异性引物进行PCR扩增,获得了1 626 bp的DNA片段。通过T-A克隆技术,将PCR产物克隆至pGEM-T载体。经鉴定,成功地构建出了重组质粒pGEM-T-hsp65。序列分析结果表明:该序列与GenBank中副结核分枝杆菌K-10株的同源性为99.2%。更多还原

【Abstract】 The genomic DNA was extracted from Mycobacterium paratuberculosis(MP) strain C-2.The secreted protein hsp65 gene was amplified with a pair of specific primers using polymerase chain reaction(PCR).PCR product was anapproximate 1 626 bp DNA segment.The clone vector pGEM-T-hsp65 was constructed successfully by the PCR product that was cloned into pGEM-T vector using T-A clone technique, and the recombinant clone was identified by using complementation test,plasmid size test,restrictional enzyme assay,plasmid PCR identification and recombinant plasmid sequence analysis.The analysis indicated that the hsp65 gene was very conservative in MP.These results could serve as a basis for further study on the usefulness of hsp65 gene and immunogenicity characteristic of hsp65 gene expression product.更多还原