结核分枝杆菌Rv0309抗原的重组表达、纯化和鉴定

Recombinant Expression, Purification and Identification of Antigen Rv0309 of Mycobacterium Tuberculosis

目的重组表达结核分枝杆菌Rv0309蛋白,以进一步探讨其免疫效应。方法将结核分枝杆菌抗原Rv0309的全长cDNA插入到原核表达载体pGEX-4T-1中,构建成Rv0309重组质粒。将重组质粒转入大肠杆菌BL21并以IPTG诱导表达,进一步经GST亲合层析柱纯化,经SDS-PAGE和Western-blot鉴定重组表达蛋白。结果获得了pGEX4T-Rv0309重组子,并在大肠埃希菌BL21中诱导表达重组Rv0309蛋白,其条带大小约23000,与预期结果相符。结论成功地进行了结核分枝杆菌抗原Rv0309的基因克隆与重组表达,为进一步研制新型结核病疫苗打下了基础。

Objective To express recombinant antigen Rv0309 of mycobacterium tuberculosis （Mtb） for the development of novel TB vaccine. Methods The whole gene containing DNA sequence encoding antigen Rv0309 was synthesized and used as a template to amplify the DNA sequence by polymerase chain reaction （PCR） for expressing recombinant Rv0309 protein, containing restriction sites on 5-and 3-end respectively. The PCR products were inserted into the expression vector pGEX-4T-1 and then the recombi- nant plasmid was transformed into E. Coli BI221, induced by isopropylthio-13-D-galactoside （IPTG）. Results The recombinant protein Rv0309 （23KD） was expressed and confirmed by SDS-PAGE and Western-blot. Conclusion The recombinant Mtb antigen Rv0309 can be expressed successfully, and it acts as a base on which new antitubereulous vaccines will be developed.