Construction method and application of recombinant adenovirus with triple epitope gene of cow mastitis pathogenic bacteria and c-type lysozyme gene of grouper

Abstract

The invention discloses a construction method and application of a recombinant adenovirus of a triple epitope gene of a cow mastitis pathogenic bacterium and a grouper c-type lysozyme gene. In the experiment, the specific protein of the main pathogenic bacteria of dairy cow mastitis was selected, and the two B cell dominant epitopes of each protein were predicted by the ABCpred scheme and the BepiPred scheme, and the obtained dominant epitopes were connected to obtain the TRE sequence, and the cLYZ sequence was obtained on GeneBank and Perform mammalian cell-preferred codon optimization. A self-cleaving peptide of Thoseaasigna virus 2A (T2A) with high shearing efficiency was inserted between the TRE sequence and the cLYZ sequence, and the pMD‑19T vector containing the target gene fragment was obtained after synthesis. Subcloning the TRE gene, T2A gene, and cLYZ gene into the shuttle vector pDC315‑MCS‑EGFP; using LipoFiter TM The recombinant shuttle vector and the adenovirus large backbone pBHGlox(delta)E1,3cre were co-transfected into HEK293 cells by liposome transfection to achieve homologous recombination and obtain the recombinant adenovirus Ad‑TRE‑T2A‑cLYZ‑EGFP.

Description

technical field [0001] The invention belongs to the field of genetic bioengineering, and relates to the prediction, design and synthesis of dominant B / T cell epitopes of Staphylococcus aureus (Staphylococcusaureus), Escherichia coli (Escherichia coli) and Streptococcus (Streptococcus), and cLYZ gene in T2A Peptide-mediated construction of a co-expression recombinant adenovirus method, specifically, a method of using bioinformatics to obtain the co-expression of the triple epitope gene (TRE) of the main pathogenic bacteria of cow mastitis and the grouper c-type lysozyme gene cLYZ The method and application of recombinant adenovirus vector construction. Background technique [0002] Cow mastitis is a serious disease that endangers the production of dairy cows and is mainly caused by the cross or co-infection of Staphylococcus aureus, Escherichia coli and Streptococcus, which has been plaguing the healthy development of the dairy industry in the world. At this stage, it mainly...

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Problems solved by technology

The frequent use of antibiotics can easily lead to problems such as bacterial resistance and drug residues, and it is not a long-term treatment method that can be relied on

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