[Cloning, expression and identification of human survivin in E. coli].
OBJECTIVE
To efficiently express and identify recombinant human survivin in E. coli.


METHODS
Survivin cDNA was amplified by reverse transcriptional (RT)-PCR and cloned into the prokaryotic expression vector pBV220, followed by expression of the recombinant plasmid in E.coli strain BL21 (Gold). To obtain survivin protein, DEAE-Sepharose Fast-Flow ion exchange chromatography and Sephacryl S-200 gel filtration were performed. Western blot analysis was used for detecting the expressed product.


RESULTS
Survivin protein was expressed in E.coli in the form of inclusion body at the expression level over 30% of the total cell protein. After ion exchange chromatography and gel filtration, the recombinant protein reached a purity over 95% and exhibited specific reaction with mouse anti-human antibody.


CONCLUSIONS
Survivin protein with high purity can be obtained by the method described above to facilitate further study of the anti-apoptosis function of survivin.
Graduate School of Kunming Medical College, Kunming 650031, China. bj520307@sina.com
Jun
J
Bie
Mao-sheng
MS
Sun
Qiang-ming
QM
Sun
Jian-ping
JP
Zheng
Wen-jia
WJ
Sun
D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D009363:Neoplasm Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D011387:Prokaryotic Cells / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D000077022:Survivin
C107489:BIRC5 protein, human; D018076:DNA, Complementary; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D000077022:Survivin