Cat omega interferon mutant as well as preparation method and application thereof
A technology of interferon and mutant, which can be used in the preparation of medicines or reagents for preventing or treating feline viral diseases, the preparation and application of cat ω interferon, and the field of cat ω interferon mutants, which can solve the problem of weak antiviral activity, The problem of low expression of cat interferon
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Embodiment 1
[0049] Embodiment 1 Expression and detection of feline interferon ω2, 9, 11 and ω2 mutant gene sequence in silkworm bioreactor
[0050] 1. Experimental method
[0051] 1.1 Acquisition, synthesis and plasmid construction of feline ω-type interferon gene
[0052] 1.1.1 Acquisition of genes
[0053] Search and obtain the gene sequence and amino acid sequence of FeIFN-ω2 (GenBank accession number: DQ420221), FeIFN-ω9 (GenBank accession number: DQ420228), FeIFN-ω11 (GenBank accession number: DQ420230.1) from NCBI.
[0054] Among them, the amino acid sequence of feline ω2 interferon is shown in SEQ ID NO.1, its gene sequence is shown in SEQ ID NO.3; the amino acid sequence of cat ω11 interferon is shown in SEQ ID NO.6.
[0055] After comparing the gene sequences and amino acid sequences of the 13 subtypes of feline ω-interferon, it was found that the amino acid composition of each subtype has very small mutations, such as amino acid changes from P to L, and G to A; cat ω-interfero...
Embodiment 2
[0073] Example 2 Expression and detection of mutated and optimized feline interferon ω11 gene in silkworm bioreactor
[0074] Based on the comparison results of Example 1, the mutation of the cat ω2 interferon gene is beneficial to the improvement of antiviral activity. Therefore, the cat ω11 type interferon gene was mutated according to the same method as ω2-M.
[0075] 1. Experimental method
[0076]1.1 Mutation, optimization and synthesis of feline ω11-type interferon gene
[0077] Obtain the gene sequence and amino acid sequence of FeIFN-ω11 (GenBank accession number: DQ420230.1) from NCBI, according to the method of Example 1, introduce 4 amino acid site-directed mutations: R82Q, P105L, V141A, G194A, and obtain 196 The amino acid mutant sequence is shown in SEQ ID NO.7, and the deduced mutant gene sequence is shown in SEQ ID NO.8. The DNAman software was used to analyze the restriction enzyme cutting sites of the mutated gene sequence. According to the analysis results ...
Embodiment 3
[0097] Embodiment 3 Mutation and optimization of cat interferon ω2 gene and expression and detection in silkworm bioreactor
[0098] Based on the results of Examples 1 and 2, cat ω-interferon gene mutation and optimization are conducive to the improvement of antiviral activity. For this reason, cat ω-2 type interferon gene and mutant gene are adjusted to ω2 and ω2-M according to the optimization method in Example 2. Gene optimization, the specific method is as follows:
[0099] 1. Experimental method
[0100] 1.1 Construction of cat ω2 interferon mutant gene and optimization and synthesis of ω2 interferon gene and mutant gene sequence
[0101] 1.1.1 Construction of cat ω2-type interferon mutant gene
[0102] The present invention is based on the comparison results of the gene sequences and amino acid sequences of 13 subtypes of feline ω-interferon in Example 1. In order to improve the expression of feline ω-interferon in the baculovirus expression system, the present inventi...
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