A visual method for direct selection of high-producing Pichia pastoris clones

文献类型： 外文期刊

第一作者： Hu, Fan

作者： Hu, Fan;Rao, Ben;Zheng, Peng;Zhou, Yu Lin;Ma, Xiang Dong;Ma, Li Xin;Li, Xin;Liu, Sheng Yi;Lue, Jie;Mao, Pei Hong;Jin, Xiang;Ke, Tao

作者机构：

期刊名称：BMC BIOTECHNOLOGY （ 影响因子：2.563； 五年影响因子：3.292 ）

ISSN： 1472-6750

年卷期： 2011 年 11 卷

页码：

收录情况： SCI

摘要： Background: The methylotrophic yeast, Pichia pastoris, offers the possibility to generate a high amount of recombinant proteins in a fast and easy way to use expression system. Being a single-celled microorganism, P. pastoris is easy to manipulate and grows rapidly on inexpensive media at high cell densities. A simple and direct method for the selection of high-producing clones can dramatically enhance the whole production process along with significant decrease in production costs. Results: A visual method for rapid selection of high-producing clones based on mannanase reporter system was developed. The study explained that it was possible to use mannanase activity as a measure of the expression level of the protein of interest. High-producing target protein clones were directly selected based on the size of hydrolysis holes in the selected plate. As an example, the target gene (9elp-hal18) was expressed and purified in Pichia pastoris using this technology. Conclusions: A novel methodology is proposed for obtaining the high-producing clones of proteins of interest, based on the mannanase reporter system. This system may be adapted to other microorganisms, such as Saccharomyces cerevisiae for the selection of clones.

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