[Establishment of prokaryotic expression and optimization ox expression conditions of Eleutherococcus senticosus P450 gene].
According to the sequence of P450 cDNA of Eleutherococcus senticosus, specific primers were designed. Frokaryotic ex pression vector pET30a-P450 was constructed and the prokaryotic expression conditions were optimized. Results showed that the BL21 after being transformed with the recombinant expression vector accumulated the high amount of recombinant protein. SDS-PAGE analysis showed that the recombinant protein was about 53 kDa. The recombinant accumulated the highest amount of recombinant protein af ter IPTG (1 mmol x L(-1)) at 27-37 degrees C for 24 h. Consequently P450 gene of E. senticosus could be expressed successfully by prokaryotic expression vector pET30a-P450. Induction temperature, IPTG concentration, medium type and amount of induction time could all influence the expression of target protein, but the impact strength was different.
Peng
P
Wu
Le-shan
LS
Xiu
Fei-fei
FF
Li
Zhao-bin
ZB
Xing
D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism; D028001:Eleutherococcus / Q000201:enzymology* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D010940:Plant Proteins; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System