title: [Expression and identification of Bm-TFF2 in Pichia pastoris].
abstract: Bm-TFF2, an amphibian trefoil factor, which is isolated from skin secretions of frog Bombina maxima, has much stronger biological activities than human TFFs. In the present study, Bm-TFF2 gene was amplified by polymerase chain reaction (PCR) from its cDNA and cloned into Pichia pastoris expression vector pPIC9K containing AOX1 promoter and alpha -factor leader sequence. Multi-copies insertion transformants were screened on G418 plates. After the induction by 1% methanol for 72 hours, the expression of Bm-TFF2 came up to the best quantity which was about 50 mg in 1L medium, and 80% saturation ammonium sulfate was suitable to collect the Bm-TFF2 protein, as identified by SDS-PAGE and Western blotting assay. The results showed that the plasmid of Bm-TFF2-pPIC9K was constructed successfully and expressed abundantly in eukaryotic expression system, which lies basis for researching further the biological activities and the relationship of structure and functions of Bm-TFF2.
authors.affiliation: Kunming Institute of Zoology, the Chinese Academy of Sciences, Kunming 650223, China.
authors.forename: Guo-Yu
authors.initials: GY
authors.lastname: Yu
authors.forename: Hong-Yun
authors.initials: HY
authors.lastname: Zhang
authors.forename: Yong
authors.initials: Y
authors.lastname: Zhang
authors.forename: Ping
authors.initials: P
authors.lastname: Jiang
authors.forename: Wen-Hui
authors.initials: WH
authors.lastname: Lee
authors.forename: Yun
authors.initials: Y
authors.lastname: Zhang
mesh_terms: D000818:Animals; D001001:Anura; D006801:Humans; D010455:Peptides / Q000235:genetics*; D010843:Pichia / Q000235:genetics*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D000071164:Trefoil Factor-2
chemical_list: D010455:Peptides; D011994:Recombinant Proteins; C000604007:TFF2 protein, human; D000071164:Trefoil Factor-2