High expression and analysis of recombinant human serum albumin in Pichia pastoris

Objective: To develop a highlevel human serum al bumin (HSA) expression system using Pichia pastoris~{!~}WT~{!=~}. Metho dsThe expression vector for HSA which was constructed in this lab was t ransformed to P.pastoris~{!~}WT~{!=~} by electroporation. The transformants wer e selected by growth on G418 and screened for HSA production by SDSPAGE and We stern blot. The HSA secretion levels were estimated by biochemical method, elect rophoresis scan and Brodford~{!d~}s method, and competitive ELISA method. Re sults The Western blot analysis showed tha t the recombinant products as a 67?000(M~{*-~}r~{!~}WT~{!=~}) secretion protein reac ted with an antiHSA monoclonal antibody. By induced expression in shake flask culture for eight days, about 3.5?g/L of rHSA was obtained from Z~{*-~}162 strain supernatant of P.pastoris. Conclusions:A yeast strai n was constructed introducing an expression vector for HSA into the P.pastoris genome. The strain prod uced as much as 3.5?g HSA per liter of shake flask culture medium.