Prokaryotic expression and purification of the cDNA of recombinant human cytokeratin 9

VernacularTitle:重组人细胞角蛋白9cDNA的原核表达及纯化
Author: Bo WANG ; Yan ZHOU ; Weikun HOU ; Yongsong CAI ; Ying ZHANG ; Linyu WANG ; Yan HAN ; Liesu MENG
Keywords: prokaryotic expression system; cytokeratin 9; fusion protein; cytoskeleton
From: Journal of Xi'an Jiaotong University(Medical Sciences) 2017;38(4):502-506
CountryChina
Language:Chinese
Abstract: Objective To clone and fuse the cDNA of human cytokeratin 9 in prokaryotic expression system,and purify and identify the fusion protein.Methods The cDNA fragment of human cytokeratin 9 was amplified from human keratinocyte (HaCaT) total RNA with specific primers.The PCR products were cloned into vector pET-28a,then the fusion protein of his-CK9 was induced by IPTG.The expressed fusion protein of his-CK9 was purified by nickel ion affinity chromatography and identified by SDS-PAGE and Western blot.Results The sequencing proved that the recombinant vector of the cDNA of CK9 was correct.The fusion protein of his-CK9 was induced to be expressed in E.coli.The fusion protein of his-CK9 was highly purified and his-CK9 showed specific binding to the commercialized antibodies of CK9.Conclusion The recombinant vector of pET-28a-CK9 has been successfully constructed,and the fusion protein of his-CK9 has been successfully expressed and purified.