Prokaryotic expression and purification of 4Aβ_(1-15)

Objective To express 4Aβ1-15 in E.coli and purify the expressed product.Methods The 1-2 Aβ1-15 gene was amplified by PCR using recombinant plasmid pcDNA3.1-4Aβ1-15 as a template,and cloned into prokaryotic expression vector pQE30a.The constructed recombinant plasmid pQE30a-2Aβ1-15 was used as a template for amplification of 2-2Aβ1-15 gene which was cloned into plasmid pQE-30a-2Aβ1-15.The constructed recombinant plasmid pQE30a-4Aβ1-15 was transformed to E.coli M15 for expression under induction of IPTG.The expressed product was identified by SDS-PAGE and Western blot,then purified by Ni2+-NTA affinity chromatography.Results Restriction analysis and sequencing proved that recombinant plasmid pQE30a-4Aβ1-15 was constructed correctly.The expressed recombinant protein,with a relative molecular mass of about 18 000,mainly existed in soluble form,contained about 40% of total protein in supernatant and showed specific binding to mouse anti-human Aβ40 monoclonal antibody,in which foreign proteins were basically removed after purification.Conclusion Recombinant prokaryotic expression vector pQE30a-4Aβ1-15 was constructed successfully,and His6-4Aβ1-15 fusion protein was expressed and purified.