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- W2373900900 abstract "Objective:To prepare full-length receptor-associated protein(RAP)and C-terminal fusion protein through the PCR and molecular cloning technology,and examine the expression of RAP1083 and RAP324 fusion protein in E.coli BL21.Methods:The renal cortex total RNA was extracted from SD rats with Trizol,and the full-length RAP cDNA was obtained by RT-PCR method.The connection of cDNA and pGEM-T cloning vector was carried out to clone and transform.The positive clones were picked and sequenced.And then the primers were designed which consist of EcoRⅠand XhoⅠdouble restriction sites according to the cDNA sequences in Gene Bank for the RAP.By running PCR,the target DNA was obtained,which aimed directly at C-terminal and the full length of RAP.The purified DNA and pGEX-4T-1 containing GST were reorganized in vitro and subcloned into competent BL21 cells.The positive clones were chosen and expanded.The fusion proteins of RAP full-length and C-terminal were prepared by IPTG induced BL21 cells,and purified by GST-Sepharose -4B affinity chromatography.Results:The sequencing results showed that the original expression vector pGEX-4T-1-RAP1083 and pGEX-4T-1-RAP324 were successfully constructed in the inducted E.coli BL21.The recombinant fusion proteins of the RAP total length of 70 ku and C-terminal 40 ku were expressed and purified.Conclusion:The prokaryotic expression vectors pGEX-4T-1-RAP1083 and pGEX-4T-1-RAP324 were constructed successfully.The purified fusion protein of GST-RAP1083 and GST-RAP324 were stably expressed in E.coli BL21,which would provide the basis for further study on the molecular pathogenesis of Heymann nephritis." @default.
- W2373900900 created "2016-06-24" @default.
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- W2373900900 date "2009-01-01" @default.
- W2373900900 modified "2023-09-24" @default.
- W2373900900 title "Role of Full-Length Fusion Protein and Receptor-Associated Protein in Heymann Nephritis" @default.
- W2373900900 hasPublicationYear "2009" @default.
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