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Article Abstract
Aim: To get BCG HSP70 protein with excellent biologic activity through E.coli. expression and purification.
Methods: BCG HSP70 gene was amplified by PCR and inserted into vector pMD18-T. After confirmed by sequencing, the gene was subcloned into expression vector pET28a. Recombinant pET28a/HSP70 was transformed into E.coli. BL21(DE3). Recombinant BCG HSP70 protein was expressed with IPTG induction and the purified protein was then identified by SDS-PAGE and Western blot. And its effect on the proliferation of mouse splenocytes was observed.
Results: Gene encoding BCG HSP70 which was identical with that published in GenBank was successfully obtained by PCR. SDS-PAGE analysis showed a protein with relative molecular mass of 70 000 was expressed. When the purified protein was detected by Western blot analysis, a specific protein with a molecular mass of 70 000 could be visualized. The purity of the purified protein was about 96.5%. The purified protein could stimulate the proliferation of mouse splenocytes significantly.
Conclusion: BCG HSP70 is expressed and purified successfully, which would lay a foundation for further research on BCG HSP70 and BCG.
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