Screening of genes of proteins interacting with p7 protein of hepatitis C virus from human liver cDNA library by yeast two-hybrid system.
OBJECTIVE
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.


CONCLUSIONS
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.
Huang
Yan-Ping
YP
Department of Pediatrics, The First Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi Province, China. huangyp423@126.com
Zhang
Shu-Lin
SL
Cheng
Jun
J
Wang
Lin
L
Guo
Jiang
J
Liu
Yan
Y
Yang
Yuan
Y
Zhang
Li-Ying
LY
Bai
Gui-Qin
GQ
Gao
Xue-Song
XS
Ji
Dong
D
Lin
Shu-Mei
SM
Zhong
Yan-Wei
YW
Shao
Qing
Q
D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics; D015723:Gene Library; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008099:Liver / Q000378:metabolism* / Q000821:virology*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D000086082:Viroporin Proteins
D004279:DNA, Viral; D011994:Recombinant Proteins; D014764:Viral Proteins; C088369:p7 protein, Hepatitis C virus; D000086082:Viroporin Proteins