Journal of Guangdong University of Technology ›› 2017, Vol. 34 ›› Issue (01): 19-23.doi: 10.12052/gdutxb.160123

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Cloning of Tenebrio Fibrinolyric Proteins gene and Expression in Pichia pastoris and Biological Activities Assay

Han Ya-li, Lin Fei-fan, Lin Ze-fei, Duan Xue-juan, Zhu Xiao-ling   

  1. School of Chemical Engineering and Light Industry, Guangdong University of Technology, Guangzhou 510006, China
  • Received:2016-09-28 Online:2017-01-09 Published:2017-01-09

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Abstract:

In this paper, TFP fragment was firstly obtained by PCR amplification using a plasmid which contains TFP cDNA as a template. The amplified fragment wascloned into P. Pastoris expressing vector pPIC9K. After confirming through sequencing, a recombinant expression plasmid pPIC9K-TFPisobtained. The plasmid islinearized and then transformed into P. Pastoris cell GS115 by electroporation. The positive recombinantis screened and confirmed by PCR. A single colony is used fermented which grew at 30℃ in a shaking incubator. TFP gene is expressed in yeast after induced with 0.5% methano and the specificity of expressed proteinidentified by SDS-PAGE. TFP fibrinolytic activity is detected by fibrin-plate method. The results showthat TFP gene has been successfully expressed in yeast Pichia pastoris, and thatthe expressed protein has fibrinolytic activity.

Key words: Tenebrio molitor L., fibrinolyric protein, gene, vector pPIC9K, Pichia pastoris GS115, expression and biological activities assay

CLC Number: 

  • R963

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