Animal genetic engineering interferon alpha and gamma composite preparations and production method and clinical application thereof

Abstract

The invention relates to an animal genetic engineering interferon compound preparation, the production method and the clinical application. The animal genetic engineering interferon compound preparation is prepared by extracting the animal peripheral blood or spleen lymphocytes, culturing, inducing by in vitro inductor, extracting the cell total RNA by Trizol, designing the specific primer, cloning the interferon gene by RT-PCR technology and connecting to the pGEM-T carrier by T-A strategy; the primer is designed again, the both ends are added with different restriction endonucleases digestion sequences, initiation codon ATG and termination codon TAA, leading peptide sequences are removed, the PCR amplification is carried out by taking the recombination T carrier as the template, so as to obtain the expression fragment of the animal interferon gene; the expression fragment is carried out with the rare codon mutation, and gel recovery target fragment is connected on the carrier of pFastBac<TM>Dual with the same double restriction endonucleases digestion after double restriction endonucleases digestion; the interferon-Alpha of an animal is cloned to multiple cloning sites under the control of Polyhedrin promoter, the interferon-Gamma of the same species animal is cloned to multiple cloning sites under the control of p10 promoter; the constructed carrier of pFastBac<TM>Dual plus interferon Alpha plus interferon Gamma are transfected to DH10Bac<TM>E.coli to carry out recombination; the constructed recombinant bacmid is extracted and purified after the blue white screening and the PCR identification, and the transfection of SF9 insect cell is carried out by a liposome method; the expression product is carried out with identification and purification; the animal genetic engineering interferon compound preparation is carried out with anti-virus activity detection and the evaluation of the clinical application effects.

Description

technical field [0001] The present invention relates to a compound preparation of animal genetic engineering interferon and its production method and application. The compound preparation of animal genetic engineering interferon is controlled by different promoters on the same carrier with animal interferon α and γ genes respectively. Interferon complex product obtained by independent expression in host cells. Background technique [0002] The outbreak of a series of livestock and poultry viral infectious diseases such as avian influenza and foot-and-mouth disease has caused huge losses to the global society and economy, directly threatening food safety and human health. In order to control and eliminate these animal diseases, the conventional way is to prevent and use antibiotics through vaccine immunization, but due to the poor breeding environment, the virus is constantly mutating. The use of antiviral chemical drugs and the strengthening of drug residue detection in foo...

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Problems solved by technology

Denaturation and enzymatic digestion not only incr