Construction and Expression of RNase-Resisting His-Tagged Virus-Like Particles Containing FluA/B mRNA.
- Author:
Jin ZHANG
;
Xiaoning XUE
;
Hefei XU
;
Ke ZHU
;
Xiaoguang CHEN
;
Juan ZHANG
;
Qi ZHANG
;
Yuan LIN
- Publication Type:Journal Article
- MeSH:
Escherichia coli;
genetics;
metabolism;
Influenza A virus;
genetics;
metabolism;
Influenza B virus;
genetics;
metabolism;
RNA, Messenger;
genetics;
metabolism;
RNA, Viral;
genetics;
metabolism;
Recombinant Fusion Proteins;
genetics;
metabolism;
Ribonucleases;
chemistry;
Virion;
genetics;
metabolism
- From:
Chinese Journal of Virology
2015;31(6):629-633
- CountryChina
- Language:Chinese
-
Abstract:
To prepare virus-like particles containing FluA/B mRNA as RNA standard and control in Influenza RNA detection, the genes coding the coat protein and maturase of E. coli bacteriophage MS2 were amplified and cloned into D-pET32a vector. Then we inserted 6 histidines to MS2 coat protein by QuikChange Site-Directed Mutagenesis Kit to construct the universal expressing vector D-pET32a-CP-His. In addition, the partial gene fragments of FluA and FluB were cloned to the down-stream of expressing vector. The recombinant plasmid D-pET32a-CP-His-FluA/B was transformed to BL21 with induction by IPTG. The virus-like particles were purified by Ni+ chromatography. The virus-like particles can be detected by RT-PCR, but not PCR. They can be conserved stably for at least 3 months at both 4 degrees C and -20 degrees C. His-tagged virus-like particles are more stable and easier to purification. It can be used as RNA standard and control in Influenza virus RNA detection.
