Cloning and Expression in Pichia pastoris of Human Granulocyte-macrophage Colony Stimulating Factor

China Biotechnology

2010, Vol. 30

Issue (01): 35-40 DOI:

Cloning and Expression in Pichia pastoris of Human Granulocyte-macrophage Colony Stimulating Factor

LI Qiao-qiao¹,WANG Qing-lu¹,ZHANG Yu-jun¹,ZHANG Rui²,XU Shou-zeng¹

1.Shandong Wanjie Medical College, Zibo 255213,China
2.Institute of Biotechnology, Chinese Academy of Agricultural Sciences,Beijing 100081,China

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Abstract

Code gene being cloned from the human genome by the primer5′extension, as the same time that code had been optimized partly to gene in the cloned procession. It was integrated into the gene secreted Pichia pastoris expression vector pPIC9K and translated yeast Pichia pastoris. PCR, SDS-PAGE and Western blotting confirmed hGM-CSF gene was integrated into the yeast genome and expression was successful. The expressed hGM-CSF was secreted into medium at the level of 389mg/L. Experiments in vivo showed that activity of hGM-CSF was normal. SDS-PAGE electrophoresis revealed that the hGM-CSF expression had occurred in glycosylation, N-glycosidase F to glycosylation also confirmed this point.

Key words： human granulocyte-macrophage colony stimulating factor primer-5 '-extension Pichia pastorion glycosylation

Received: 21 September 2009 Published: 27 January 2010

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	LI Qiao-Qiao
	WANG Qing-Lu
	ZHANG Yu-Jun
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LI Qiao-Qiao, WANG Qing-Lu, ZHANG Yu-Jun, ZHANG Dui, XU Shou-Ceng. Cloning and Expression in Pichia pastoris of Human Granulocyte-macrophage Colony Stimulating Factor. China Biotechnology, 2010, 30(01): 35-40.

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https://manu60.magtech.com.cn/biotech/ OR https://manu60.magtech.com.cn/biotech/Y2010/V30/I01/35

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