利用酵母双杂交技术筛选肝再生增强因子结合蛋白

Screening of augmenter of liver regeneration-binding proteins by yeast-two hybrid technique.

作者信息

Cheng Jun, Wang Lin, Li Ke, Lu Yin-Ying, Wang Gang, Liu Yan, Zhong Yan-Wei, Duan Hui-Juan, Hong Yuan, Li Li, Zhang Ling-Xia, Chen Ju-Mei

机构信息

Gene Therapy Research Center, Institute of Infectious Diseases, 302 Hospital of PLA, Beijing 100039, China.

出版信息

Hepatobiliary Pancreat Dis Int. 2003 Feb;2(1):81-4.

PMID:14607653

Abstract

OBJECTIVE

To investigate the biological function of augmenter of liver regeneration (ALR), we used yeast-two hybrid technique to detect proteins in hepatocytes interacting with ALR.

METHODS

ALR bait plasmid was constructed by using yeast-two hybrid system 3, then transformed into yeast AH109. The transformed yeast was mated with yeast Y187 containing liver cDNA library plasmid in a 2XYPDA medium. Diploid yeast was plated on a synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal for selection and screening. After extracting and sequencing of the plasmid from blue colonies. Analysis was performed by bioinformatics.

RESULTS

Of 36 colonies sequenced, 14 are metallothionein, 12 albumin, and 3 selenoprotein P. One colony is a new gene with unknown function.

CONCLUSION

The successful cloning of gene of ALR interacting protein has paved the way for studying the physiological function of ALR and associated proteins.

摘要

目的

为研究肝再生增强因子（ALR）的生物学功能，我们采用酵母双杂交技术检测肝细胞中与ALR相互作用的蛋白质。

方法

利用酵母双杂交系统3构建ALR诱饵质粒，然后转化至酵母AH109。将转化后的酵母与含有肝脏cDNA文库质粒的酵母Y187在2XYPDA培养基中进行杂交。将二倍体酵母接种在含有x-α-半乳糖的合成缺陷营养培养基（SD/-Trp-Leu-His-Ade）上进行筛选。从蓝色菌落中提取质粒并测序后，通过生物信息学进行分析。