Bovine rotavirus recombinant VP7 protein antigen and preparation method thereof

Abstract

The invention provides a bovine rotavirus recombinant VP7 protein antigen and a preparation method thereof and relates to a protein antigen and the preparation method thereof, and the bovine rotavirus recombinant VP7 protein antigen can be used for solving the problems that the existing bovine rotavirus detection method cannot simultaneously meet the requirements of equipment and operation simplification, good specificity and large-scale clinical popularization. The bovine rotavirus recombinant VP7 protein antigen is coded by 644bp basic groups and has a size of 40kDa. The preparation method comprises the following steps: 1, extracting genome RNA of a bovine rotavirus BRV-DQ strain; 2, carrying out reverse transcription, PCR (Polymerase Chain Reaction) amplification and purification on the genome RNA and then connecting the genome RNA with a pMD18-T carrier so as to obtain pMD18-T-VP7 plasmids; 3, simultaneously carrying out enzyme digestion on pMD18-T-VP7 and pET30a, connecting and converting into host bacteria so as to obtain recombinant bacteria pET30a-VP7 / BL21; 4, carrying out inducible expression on recombinant protein; and 5, purifying the recombinant protein so as to obtain the bovine rotavirus recombinant VP7 protein antigen. The bovine rotavirus recombinant VP7 protein antigen is applied to the field of detection of bovine rotaviruses.

Description

technical field [0001] The invention relates to a protein antigen and a preparation method thereof. Background technique [0002] Bovine rotavirus (Rotavirus, RV) belongs to the family Reoviridae and is one of the main pathogens of non-bacterial diarrhea in calves. Bovine rotavirus was first discovered by Mebus et al. in 1968 and proved to be the pathogen of neonatal calf diarrhea, named NCDV strain (NebraskacalfdiarrheaVirus, NCDV). The virus can cause depression, watery diarrhea, acidosis and loss of appetite in animals, mostly infecting calves aged 15-45 days. In 1980, for the first time in my country, rotavirus was detected in buffalo and cattle diarrhea feces, proving the existence of the virus in our country. Subsequent reports of viral diarrhea in calves followed. Bovine rotavirus has become a worldwide disease because of its wide prevalence, high incidence and great harm to calves. If secondary to other diseases, the mortality rate will increase greatly, causing ...

AI Technical Summary

Problems solved by technology

Its deficiency is mainly due to the fact that the RNA of the virus is easily degraded by RNase in the air or medicines, so that the results are inaccurate and false negatives occur; the classic PCR detection method for BRV is reverse transcription PCR (RT-PCR), bovine group A rotavirus The two nucleotide sequences that are conserved in all types of its structural protein VP7 are often used as primers to quickly and specifically identify viruses, but because the RNA extraction process is complicated and easy to contaminate, it is more suitable for laboratory diagnosis. Large-scale clinical popularization is still difficult; serological detection method is undoubtedly an ideal diagnostic method in clinical detection, among which ELISA diagnostic method does not require expensive equipment, low cost, high sensitivity and specificity, and no radioactive pollution. It is easy to promote at the grassroots level, and the indirect ELISA method established by expressing the capsid protein has the advantage of overcoming the potential risks of antigen detoxification and detoxification

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