Construction of Recombinant Mice Factor Ⅶ-pPIC9K Vector and Expression in Pichia / 中国肿瘤生物治疗杂志

ABSTRACT

Objective:To construct the yeast expressive vector of rmFⅦ, in which mFⅦ was mutated to inhibit coagulation without affecting the affinity for TF, and express it in Pichia pastoris. Methods:The full length cDNA encoding mFⅦ was amplified from a mouse liver by RT-PCR method, site-direct mutated and restriction enzyme digested as design. Cloning into pPIC9K, electroporation of Gs115, in vivo screen of multiple inserts by G418 resistance, BMGY/BMMY are used for induction and expression of rmFⅦ in pichia pastoris. These proteins were also screened for functional activity. Results: Three different rmFⅦ-pPIC9K yeast expression vectors and it's aim protein were obtained,two kinds of proteins were found to be functional active as design. Conclusion:rmFⅦ protein can be expressed in pichia pastoris and it might facilitate the development of tumor-target molecule, and novel anti-agiogenesis drug study.