Method for efficiently expressing canine circovirus (CanineCV) Cap protein

Abstract

The invention discloses a method for efficiently expressing canine circovirus (CanineCV) Cap protein. The method comprises the steps of: adopting a CanineCV genome as a template sequence, designing specific primers, performing PCR amplification so as to obtain a complete sequence of the Cap protein, intercepting a nuclear localization signal peptide which comprises 26 amino acids at the N-terminalof the Cap protein through online prediction of nuclear localization signals of the Cap protein, connecting the Cap protein sequence which is obtained after interception is conducted to a pET-28a vector so as to obtain a recombinant plasmid pET(28a)-Cap, transferring the recombinant plasmid into a BL21(DE3) competent cell, and performing induced expression by using IPTG, wherein it is shown by SDS-PAGE analysis that the recombinant Cap protein is correctly expressed in Escherichia coli BL21, so that the purpose of efficient expression of the CanineCV Cap protein is achieved. Through the method, the difficult problems of no expression or low expression level of the CanineCV Cap protein in the Escherichia coli can be solved successfully; the expressed protein has good immunogenicity, and the method can be applied to establishment of an epidemiological diagnostic method and researches of subunit vaccine of the Cap protein.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and in particular relates to a method for efficiently expressing Cap protein of canine circovirus (CanineCV) lacking 26 nuclear localization signals by using pET-28a expression vector. Background technique [0002] Canine circovirus (Canine circovirus, CanineCV) belongs to the genus Circovirus (Circoviridae), and is a single-stranded circular DNA virus with icosahedral symmetry and no envelope. The genome length of CaineCV is 2063bp, including two main open reading frames, encoding replicase Rep protein and nucleocapsid Cap protein respectively. After CaineCV was first reported in 2012, it was discovered in Italy, Germany and China. Li et al detected CaineCV in the lymph nodes and spleens of multiple dogs with vascular injury or histiocyte inflammation. Decaro et al. reported the detection of CanineCV in dead puppies from an Italian kennel that had an outbreak of acute enteritis. CanineCV ha...

AI Technical Summary

Problems solved by technology

In 2016, Sun Wenchao et al. conducted the first survey on the prevalence of CaineCV in my country. The results showed that the CanineCV strains popular in China and the strains popular in Europe and the United States are in two different branches. At present, there is no progress in the research on CaineCV. The sex and pathogenic mechanism are still unclear, and there is no ELISA detection kit and neutralizing antibody for this virus, which brings great difficulties to clinical prevention and control

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