关键词: P21活化激酶, 截短区域, 原核表达质粒, GST融合蛋白

Abstract: Abstract:Objective
To construct the prokaryotic expression plasmids of   truncated regions of human  p21-activated kinase 6(PAK6) to induce and identify their recombinant proteins,and to provide basis for discussing the biological function of PAK6 gene.Methods The eukaryotic expression plasmid pcDNA3.1-GFP-PAK6  was used as the template,and the truncated segments  of PAK6 gene were amplified by PCR method. The truncated segments were cloned into GST-tagged prokaryotic expression vector pGEX-5X-1 after do uble enzyme digestion of  EcoRⅠ/XhoⅠ.The truncated plasmids were transfect
ed into E.coli BL21 and the  PAK6 gene truncated fusion proteins were induced by IPTG and verified by Western blotting method.Results The vector fragment (5 000 bp) and PAK6 1-55（165 bp），PAK6 56-210（465 bp），PAK6 211-410（600 bp），and PAK6 385-681（891 bp）  vector fragments being consistent with the expected fragments were obtained after  double enzyme digestion of EcoRⅠ/XhoⅠ.The Western blotting  results showed that the relative molecular mass of GST-PAK6 truncated fusion proteins of PAK6 truncated region plasmids were 32 000,43 000,48 000,and 60 000,respectively.
Conclusion GST-tagged prokaryotic expression plasmids of different truncated regions of  PAK6 gene are constructed and their recombinant proteins are expressed successfully.

Key words: p21-activated kinase 6, truncated region, prokaryotic expression plasmid, GST fusion protein