Biotechnology Bulletin ›› 2013, Vol. 0 ›› Issue (10): 148-152.

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Constuction the Pichia pastoris Expression System of Neuritin and Its Impacts on PC12 Cell

Zhang Shujun1,2,, Zhao Chen3, Di Jianjun1,2,, Mushamoli1,2,, Xian Lingling4, Yu Na4, Huang Jin4,   

  1. （1. College of Life Science，Inner Mongolia Uniity for Nationalities，Tongliao 028000 ；2. Inner Mongolia Industril Engineering Research Center of Universities for Castor，Tongliao 028vers000 ；3. Department of General Surgery，Baotou Northern Hospital，aBaotou 014000 ； 4. Key Laboratory of Xinjiang Endemic and Ethnic Disease University，Shihezi 832002）
  • Received:2013-03-27 Revised:2013-10-15 Online:2013-10-14 Published:2013-10-15

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Abstract:

It was to intend to construct Pichia pastoris expression system of neuritin, to obtain large amount of active Neuritin protein and study its neurobiological function. Encoding cDNA sequence of neuritin was amplified by PCR, then the fragment was digested and ligated into shuttle vector pPIC9K. The neuritin-pPIC9K recombinant was transformed into E. coli DH5α, and was confirmed by PCR and sequencing. The recombinant was linearized by restriction enzyme Sal Ⅰ , and transformed into yeast GS115 by electric shock. After screening and identification, the Pichia pastoris expression system of neuritin was successfully constructed. Using methanol to induce the expression of neuritin, through SDSPAGE and Western blot analysis, we obtained Neuritin protein about 11 kD. The purified Neuritin protein was added into PC12 cell culture medium and it can promote the neurite growth of PC12 cell.

Key words: Neuritin pPIC9K, Pichia pastoris expression system, PC12 cell