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- W2368307744 abstract "Objective To construct a prokaryotic expression vector so that the maximum level of homogeneous,soluble,and fully bioactive recombinant human survivn/Trx fusion protein can be obtained from gastric cancer.Methods Total RNAs were extracted from human gastric cancer cell line and the survivin gene was fished by reverse transcription PCR; the survivin gene was amplified by a pair of special primer including endonucleases sites of BamH I and HindⅢby PCR and inserted into the prokaryotic expression vector pET 32a(+).After being selected by Ampicillin resistance,identified by PCR and double digestion of endonucleases,the sequenced DNA of survivin was analyzed by BLAST.The survivin/Trx fusion protein was expressed in gastric cancer.BL21(DE3)was induced with IPTG,identified by western blot and purified with Ni-NTA agarose,respectively.Results Through RT-PCR and 1% agarose gel electrophoresis, the products of 440 bp were obtained as expected.After double digestion with endonucleases,the survivin gene was inserted into the prokaryotic expression vector pET 32a(+).DNA sequencing indicated that the constructed expression plasmid was transformed into competent cells E.coli BL21(DE3).The recombinant survivin/Trx fusion protein was specific to survivin antibody identified by Western-Blotting.The purity of the protein was over 80% after being purified by Ni-NTA agarose.Conclusion Prokaryotic expression plasmid PET 32a(+)/survivin was successfully constructed and highly purified survivin/Trx fusion protein was obtained." @default.
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- W2368307744 date "2007-01-01" @default.
- W2368307744 modified "2023-09-25" @default.
- W2368307744 title "Expression and purification of survivin protein" @default.
- W2368307744 hasPublicationYear "2007" @default.
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