关键词: 人促黄体激素释放激素, 绿脓杆菌外毒素A, 可溶性表达, 表达菌株

Abstract:

We obtained fulllength nucleic acid sequence of an recombinant immunotoxin of GnRH and PE39KDEL by gene synthesis based on the E. coli codon bias. Factor Xa restriction site was inserted at the C termanol of GnRHPE39KDEL to obtion pET24bGnRHPE39KDEL recombinated plasmid which was then converted into E. coli BL21 (DE3) and BL21 (DE3) plyS. Results show that recombinant immunotoxin exhibits highlevel soluble expression after 0.2 mmol/L IPTG induction at 30 ℃ for 2 h in LB media with 0.2 % glucose till OD600 is acquired at 37 ℃ for about 0.6 h. The purity of fusion protein is above 95% and the protein yield is about 2.6 mg/g after sonication, highspeed centrifugation, HisTrap column purification and desalt. IC-50 detection indicates that the recombinant protein can significantly inhibit the growth of tumor cell strains.

Key words: GnRH, PE39KDEL, soluble expression, expression strain