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《Journal of Guangxi Normal University(Natural Science Edition)》 2016-01
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Prokaryotic Expression Optimization,Point Mutation Correction and Cloning of Human Interleukin-24

FAN Qiuli;SONG Degui;College of Life Science,Guangxi Normal University;  
Taking the interleukin(IL-24)in the lab as the template,recombinant prokaryotic vector of expressing pET28a-IL-24 was constructed,the vector was transferred into E.coli DH5α and the monoclones was picked out.The mutant(G→A in IL-24cDNA)resulted in the change of IL-24protein(A412T),The mutants(A→G,C→T,in IL-24cDNA)did not change the amino acids on the bit.By designing mutation primers,correction of mutation was performed by a two-step PCR reaction,the correct recombinant vector was transformed into E.coli.BL21.The optimal condition was confirmed by changing the concentration of IPTG and induction time and temperature,the fusion protein 6his-IL-24 was detected by SDS-PAGE and Western blotting.As a result,by two-step PCR,the prokaryotic expression vector was constructed,the expression product was existed in the form of inclusion body,the best induction temperature was 28 ℃,the optimal IPTG concentration was 1 mmol/L and the best induction time was 8h.
【Fund】: “桂科重”研究项目(14121003-3-1)
【CateGory Index】: Q78
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