Construction and Identification of the Bait Vector Containing Duck Circovirus Cap Gene for the Yeast Two-hybrid System.
- Author:
Yu XU
;
Zhilong ZHANG
;
Yanyan LU
;
Lei ZHANG
;
Pengfei LI
;
Renyong JIA
- Publication Type:Journal Article
- MeSH:
Animals;
Capsid Proteins;
genetics;
metabolism;
Circovirus;
classification;
genetics;
isolation & purification;
Cloning, Molecular;
Ducks;
Genetic Vectors;
genetics;
metabolism;
Recombinant Fusion Proteins;
genetics;
metabolism;
Saccharomyces cerevisiae;
genetics;
metabolism;
Two-Hybrid System Techniques
- From:
Chinese Journal of Virology
2015;31(3):282-286
- CountryChina
- Language:Chinese
-
Abstract:
To construct a bait expression vector containing the duck circovirus Cap gene for use in the yeast two-hybrid system, the whole cap codon-optimized gene was inserted into pGBKT7 vector and confirmed by PCR, restriction enzyme digestion, and sequence analysis. After transformation into a Y2HGold yeast strain, the expression of Cap protein was analyzed by Western blotting. Toxicity and self-activation of the bait protein were detected using different dropout minimal base. PCR reaction, restriction enzyme digestion, and sequencing analyses indicated that the duck circovirus Cap gene was correctly inserted into pG- BKT7. Western blotting showed that the whole Cap protein was expressed. The recombinant bait protein had no toxicity and self-activation. Therefore, the bait vector with the Cap gene was constructed successfully, providing a foundation for future screening for interacting proteins in the yeast two-hybrid system.
