Expression of Trehalose Synthase in Pichia pastoris

The expression vector was constructed from trehalose synthase(TreS) gene(AE015451.1)of Pseudomonas putida to express in Pichia pastoris. TreS gene was amplified by PCR and double digested by EcoRⅠ/XbaⅠ, then linked to the pPICZaA which digested with same enzymes. The gene was identified, sequenced and transformed into pichia pastoris GS115 by electroporation. Positive colonies harboring target genes were screened out by the YPD medium with Zeocin. The genome of positive colonies were extracted and a same size band with target gene was abtained by using PCR which illustrated target gene had successfully transferred to Pichia pastoris. SDS-PAGE and HPLC results showed that the recombinant enzyme had a molecular of 76 ku band which was consistent with the predicted molecular mass, and it had the ability to catalyze the conversion of maltose into trehalose. The recombinant pPICZaA-TreS plasmid successfully constructed and integrated into Pichia pastoris genome and expressed as expected.