Screening of hepatocyte proteins binding with the middle surface protein of the hepatitis B virus by the yeast two‑hybrid system
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- Published online on: March 24, 2014 https://doi.org/10.3892/mmr.2014.2069
- Pages: 2342-2346
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Abstract
The effect of the middle hepatitis B virus surface protein (MHBs) remains to be elucidated. To investigate the biological function of the MHBs protein, the present study performed yeast two‑hybrid screening to search for proteins that interact with the MHBs protein in hepatocytes. The bait plasmid expressing the MHBs protein was constructed by cloning the gene of the MHBs protein into pGBKT7, then the recombinant plasmid DNA was transformed into AH109 yeast (a type). The transformed yeast AH109 was mated with yeast Y187 (α type) containing the liver cDNA library plasmid in 2X yeast peptone dextrose adenine (YPDA) medium. The mated diploid yeast was plated on quadruple dropout medium (SD/‑Trp‑Leu‑His‑Ade) containing X‑α‑gal for selection and screening. Following extracting and sequencing of the plasmids from positive (blue) colonies, the sequence analysis was conducted and analyzed by bioinformatics methods. Two colonies were selected and sequenced. Among them, one was the human DNA sequence from the clone RP11‑490D19 on chromosome 9 and the other was homo sapiens 12 BAC RP11‑180M15 (Roswell Park Cancer Institute Human BAC Library). The yeast two‑hybrid system is an effective method for identifying hepatocyte proteins that interact with MHBs. The MHBs protein binds with different proteins suggesting that it has multiple functions in vivo.
