Detection method for biological activity of porcine interferon alpha through luciferase reporter gene method

• Main Authors: LI YASEN, XU GAOTAO, JIANG MINZHI, SHAN XUEQIN, LIU JIALU, ZHAO YU, FAN YUFANG
• Format: Patent
• Language: Chinese; English
• Published: 26.10.2018
• Subjects: BEER; BIOCHEMISTRY; CHEMISTRY; COMPOSITIONS OR TEST PAPERS THEREFOR; COMPOSITIONS THEREOF; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL ORENZYMOLOGICAL PROCESSES; CULTURE MEDIA; ENZYMOLOGY; INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIRCHEMICAL OR PHYSICAL PROPERTIES; MEASURING; MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEICACIDS OR MICROORGANISMS; METALLURGY; MICROBIOLOGY; MICROORGANISMS OR ENZYMES; MUTATION OR GENETIC ENGINEERING; PHYSICS; PROCESSES OF PREPARING SUCH COMPOSITIONS; PROPAGATING, PRESERVING OR MAINTAINING MICROORGANISMS; SPIRITS; TESTING; VINEGAR; WINE

The invention discloses a detection method for the biological activity of a porcine interferon alpha through a luciferase reporter gene method and application of the detection method. The method comprises the following steps: obtaining a gene segment of pMx1 of porcine Mx1 protein by adopting PCR (Polymerase Chain Reaction) amplification; inserting the gene segment of the pMx1 into a 5' end of a pGL3-basic vector luc gene; then carrying out the PCR amplification to obtain a pMx1-luc fused gene segment; replacing gene segments of pCMV (Porcine Cytomegalovirus) and EGFP (Enhanced Green Fluorescent Protein) in a pEGFP-N1 vector by utilizing the pMx1-luc fused gene segment; constructing a pMx1-luc plasmid and transfecting a cell; selecting a stably-transfected cell line; carrying out clonal culture on the screened stably-transfected cell line; preparing a standard curve by taking a porcine interferon alpha standard product which is subjected to gradient dilution; adding a porcine interferon alpha sample to be det