Construction of α-galactosidase pYD1-84411 Recombinant Plasmid

Rice α-galactosidase(BAC84411) recombinant plasmid pET32a+-84411 as template, PCR amplification conditions and the reaction system were screened. The amplified fragment were detected by agarose gel electrophoresis, a clear and single band of the target gene was obtained. And the gene was inserted into the yeast surface display vector pYD1 by double restriction endonuclease. the insert and the vector ligased by moles ratio of 4 ∶ 1. Then the ligation mixture was transformed into E.coli JM109, and the positive recombinant plasmid was identified. The obtained pYD1- 84411plasmid would be used for inducing α-Galactosidase expression in yeast strains EBY100.