Screening of genes of proteins interacting with p7 protein of hepatitis C virus from human liver cDNA library by yeast two-hybrid system
Screening of genes of proteins interacting with p7 protein of hepatitis C virus from human liver cDNA library by yeast two-hybrid system
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DOI：
10.3748/wjg.v11.i30.4709
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发表时间：
2005-08-14
影响因子：
4.3
通讯作者：
Shao, Qing
中科院分区：
文献类型：
--
作者：
Huang, Yan-Ping;Zhang, Shu-Lin;Shao, Qing
目的：研究p7蛋白的生物学功能，寻找肝细胞内与p7蛋白相互作用的蛋白。方法：将p7蛋白基因克隆至pGBKT7中，构建p7蛋白诱饵质粒，转化至酵母AH109（一型）中。将转化的酵母与含有肝cDNA文库质粒pACT2的酵母Y187（α型）在2xYPDA培养基中交配。将二倍体酵母铺在含有 x-α-gal 的合成脱落营养培养基 (SD/-Trp-Leu-His-Ade) 上进行选择和筛选。蓝色菌落提取质粒并测序后，通过生物信息学进行序列分析。结果：筛选出50个菌落并测序。其中，1个集落为智人信号序列受体，7个集落为智人H19，7个集落为含有富含亮氨酸重复序列的免疫球蛋白超家族，3个集落为精细胞核周RNA结合蛋白，2个集落为跨膜4结构域，24个集落为癌症相关抗原，4个集落为核孔蛋白214 ku，2个集落为CLL相关结论：p7蛋白相互作用蛋白基因的成功克隆，为p7蛋白及其相关蛋白的生理功能研究奠定了基础。 (C) 2005 The WJG Press 和 Elsevier Inc. 保留所有权利。
AIM: To investigate the biological function of p7 protein and to look for proteins interacting with p7 protein in hepatocytes.METHODS: We constructed p7 protein bait plasmid by cloning the gene of p7 protein into pGBKT7, then transformed it into yeast AH109 (a type). The transformed yeast was mated with yeast Y187 (alpha type) containing liver cDNA library plasmid, pACT2 in 2xYPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal for selection and screening. After extracting and sequencing of plasmids from blue colonies, we performed sequence analysis by bioinformatics.RESULTS: Fifty colonies were selected and sequenced. Among them, one colony was Homo sapiens signal sequence receptor, seven colonies were Homo sapiens H19, seven colonies were immunoglobulin superfamily containing leucine-rich repeat, three colonies were spermatid peri-nuclear RNA binding proteins, two colonies were membrane-spanning 4-domains, 24 colonies were cancer-associated antigens, four colonies were nucleoporin 214 ku and two colonies were CLL-associated antigens.CONCLUSION: The successful cloning of gene of protein interacting with p7 protein paves a way for the study of the physiological function of p7 protein and its associated protein. (C) 2005 The WJG Press and Elsevier Inc. All rights reserved.