Method of detecting biological activity of bovine interferon alpha by luciferase reporter gene process

A biological activity and reporter gene technology, applied in the field of interferon activity detection, can solve the problems of inconvenience for large-throughput sample detection, hidden dangers of live virus biological safety, and unsatisfactory sensitivity, so as to avoid biological safety hazards and shorten detection. time, the effect of improving repeatability
CN108753918AInactive Publication Date: 2018-11-06ANHUI JIUCHUAN BIOTECH
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
ANHUI JIUCHUAN BIOTECH
Publication Date
2018-11-06
Estimated Expiration
Not applicable · inactive patent

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Abstract

The invention discloses a method of detecting biological activity of bovine interferon alpha by luciferase reporter gene process and application of the method. The method comprises the steps of performing PCR (polymerase chain reaction) amplification to obtain a gene fragment of pMx1 of bovine Mx1 protein; inserting the gene fragment to 5' end of pGL3-basic vector luc gene; performing PCR amplification to obtain a pMx1-luc fusion gene fragment; replacing gene fragments of pCMV and EGFP in pEGFP-N1 vector via the pMx1-luc fusion gene fragment so as to construct pMx1-luc plasmid, transfecting cells, and selecting a cell strain that is stably transfected; subjecting the screened cell strain that is stably transfected to cloning culture; using bovine interferon alpha standard gradiently diluted to prepare a standard curve, adding a bovine interferon alpha sample under detection into the cells that are subjected to cloning culture, allowing co-incubation, detecting fluorescence intensity, and evaluating titer of the bovine interferon alpha sample under detection through the standard curve.
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Description

technical field

[0001] The invention relates to a method for detecting the biological activity of bovine interferon alpha by luciferase reporter gene method, belonging to the technical field of interferon activity detection. Background technique

[0002] Interferon α is widely used in the treatment of diseases in antiviral infection and immune dysfunction. As a cytokine, it can activate a series of intracellular signal transduction pathways by binding to specific receptors indicated by target cells, showing antiviral effects. or immunomodulatory effects. At present, the signal transduction pathway of interferon α in cells has been studied thoroughly, mainly through the activation of JAK-STAT signal cascade. Interferon α binds to receptors to activate receptor-related JAK1 and TYK2, phosphorylate STAT1 and STAT2 tyrosine, phosphorylated STAT1 and STAT2 form dimers and transfer into the nucleus, and assemble interferon regulatory factor 9 (IRF9) to form 3-mer interferon-stim...

Claims

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