Title: Cloning and Expression of Human Heat Shock Protein 70 Gene and Identification of Expressed Product
Abstract: Objective To clone human heat shock protein 70(HSP70)gene for the construction of a prokaryotic expression vector.Methods Amplify HSP70 gene by PCR,clone into prokaryotic expression vector pET-28b,then transform to E.coli JM109.Screen the positive clones,extract plasmid pE28b70F and transform to E.coli BL21(DE3)for expression under induction of IPTG.Results The expressed HSP70 contained 22.3% of total somatic protein,of which 12% were soluble protein.Western blot showed specific reaction of the expressed protein with mouse anti-human HSP70 McAb.About 15-20 mg of recombinant protein was purified from 1 L of culture supernatant of recombinant E.coli by Ni~2+-NTA affinity column chromatography,and reached a purity of more than 80%.Conclusion HSP70 was successfully expressed,which laid a foundation of further study on its biological function and action mechanism.
Publication Year: 2007
Publication Date: 2007-01-01
Language: en
Type: article
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