[Screening and cloning of the genes of protein interacting with the N-terminal protein of hepatitis B virus DNA polymerase by yeast-two hybrid technique].
OBJECTIVE
To screen and clone the genes of protein interacting with the N-terminal protein (TP) of hepatitis B virus DNA polymerase.


METHODS
TP was amplified by polymerase chain reaction (PCR) and TP bait plasmid was constructed by using yeast two-hybrid system 3, then transformed into yeast AH 109. The transformed yeast was mated with yeast Y187 containing liver cDNA library plasmid in 2 x YPDA medium. Diploid yeast was plated on synthetic dropout medium (SD/-Trp-Leu-His-Ade) and that containing X-alpha-GAL for selecting two times and screening. Plasmids were extracted from blue colonies, and sequence analysis was performed by bioinformatics.


RESULTS
Forty-seven clones were obtained, these clones included human P36956 sterol regulatory element binding protein-1, RNA polymerase II subunit hsRPB7 mRNA, asialoglycoprotein receptor 2, transcript variant 3, ceruloplasmin (ferroxidase), transmembrane 4 superfamily member 2 and 19 of the hypothetical proteins and so on.


CONCLUSIONS
Genes encoding TP interacting proteins in hepatocytes were successfully cloned and the results suggest that TP has a wide variety of biological functions.
Chen
Guo-feng
GF
The 4th Department, The No 302 Hospital of The People's Liberation Army, Beijing 100039, China.
Wang
Lin
L
Cheng
Jun
J
Zhang
Ling-xia
LX
Li
Li
L
Zhang
Jian
J
Shao
Qing
Q
Ji
Dong
D
D003001:Cloning, Molecular; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015723:Gene Library; D006515:Hepatitis B virus / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D008099:Liver / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D011991:Receptors, Virus / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D014170:Transformation, Genetic; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D011991:Receptors, Virus; D014764:Viral Proteins; C438016:hepatitis B virus-binding protein, human; D004259:DNA-Directed DNA Polymerase