Title: PROKARYOTIC EXPRESSION OF RECOMBINANT MOUSE OP-1 AND ITS PURIFICATION
Abstract: Objective To investigate the effective means of rmOP-1 cloning, construction of the prokaryotic expression vector, and its purification. Methods mRNAs were isolated and purified from a 2-week embryo of a CD-1 mouse. cDNAs were synthesized by Oligo(dT) primers with AMV. Full-length open reading frame of mOP-1 was screened by PCR. PCR products was inserted into pCR 3.1-Uni vector by TA cloning. Subcloned into pTrcHis 2B and induced by IPTG, recombinant OP-1 was expressed with prokaryotic expression system, E. coli, JM109. The recombinant protein was isolated and purified with Ni-NTA resin, and its osteogenesis was assayed in an ALP activity. Results Open reading frame was verified by both restrictive endonuleotidase analysis and DNA sequencing. rmOP-1 purified from Ni-NTA resin chromatography was analyzed by PAGE. Conclusion Over-expression of OP-1 mRNA exists in a 2-week embryo of a CD-1 mouse. The gene can be easily obtained by RT-PCR and TA cloning. The high efficiency of prokaryotic expression vector pTrcHis 2B would offer a fast and simple means of rmOP-1 production.
Publication Year: 2004
Publication Date: 2004-01-01
Language: en
Type: article
Access and Citation
AI Researcher Chatbot
Get quick answers to your questions about the article from our AI researcher chatbot