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A kind of lentiviral vector capable of directly responding to type I interferon response and its preparation method and application

A technology of lentiviral vector and interferon, which is applied in the field of medicine, can solve the problems of inappropriateness, inability to specifically and accurately respond to interferon regulation mechanism, and inability to respond well

Active Publication Date: 2018-05-01
FANTASIA BIOPHARMA ZHEJIANG CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, ISRE-Luc cannot specifically and accurately reflect the regulatory mechanism of intracellular interferon; secondly, the secondary amplification by ISRE needs to depend on the interferon recognition receptors of cells (IFNAR1, etc.), therefore, it is not suitable for knocking out IFNAR1 Or detect the interferon regulation mechanism of the cell under the condition of STAT1 and other activities being affected; third, ISRE-Luc requires transcription and translation levels, which cannot respond well to some molecules or drugs at the transcription level or translation regulation of interferon response

Method used

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  • A kind of lentiviral vector capable of directly responding to type I interferon response and its preparation method and application
  • A kind of lentiviral vector capable of directly responding to type I interferon response and its preparation method and application
  • A kind of lentiviral vector capable of directly responding to type I interferon response and its preparation method and application

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Embodiment Construction

[0039] The invention specifically discloses a sensitive and specific IRF3-BiLC reporter cell line that directly responds to type I interferon response and a construction method. It specifically includes:

[0040] 1. Construction of IRF3-BiLC lentiviral expression vector

[0041] The principle of BiLC is: cut Gaussia luciferase at certain specific sites to form two polypeptides at the N and C terminals without luciferase activity, called N-fragment and C-terminal Fragment (C-fragment) (Remy and Michnick, 2006; Cassonnet et al., 2011; Tannous et al., 2005). The two fragments cannot spontaneously assemble into an active luciferase protein when co-expressed in cells or mixed in vitro. However, when the fragments of the two luciferase proteins are respectively linked to a set of interacting target proteins, and the two fusion proteins are co-expressed in cells or mixed in vitro, due to the interaction of the target proteins, the luciferase The two fragments of the protein are cl...

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Abstract

The present invention provides a lentiviral vector capable of directly responding to type I interferon responses. The full-length pDonor-IRF3 shuttle plasmid is homologously recombined and cloned into a lentiviral BiLC expression vector through cloning technology; the lentiviral BiLC expression vector Gaussian luciferase is divided into N-terminal and C-terminal from the 109th amino acid position, denoted as GlucN and GlucC, forming a lentiviral expression vector IRF3‑BiLC. Beneficial effects of the present invention: First: Compared with the type I interferon response reporting system currently used, the reporter system of IRF3-BiLC has a specific response to type I interferon. Second: The reporter system of IRF3-BiLC can directly and rapidly respond to transient or persistent type I interferon responses.

Description

technical field [0001] The invention relates to a lentiviral vector capable of directly responding to type I interferon response, a preparation method and application thereof, and belongs to the technical field of medicine. Background technique [0002] When the human body is infected by viruses such as influenza virus, hepatitis C virus and herpes virus, etc., as well as bacterial infections, it will trigger innate immunity and adaptive immunity (1). Among them, the innate immune response mainly induces the production of interferon (Interferon, IFN) and activates downstream interferon-induced genes (ISGs), thereby exerting antiviral effects (2,3). For RNA viruses, such as influenza virus and hepatitis C virus, the virus infects and enters cells, and the released RNA can be recognized by intracellular RNA receptors such as RIG-I, MDA5, etc., and then recruit IPS-1 protein on mitochondria and phosphorylate TBK1 protein, phosphorylated TBK1 protein can cause phosphorylation o...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/867C12N15/65
CPCC12N15/65C12N15/86C12N2800/107C12N2740/15043C12N2740/16043C12Q1/66C07H21/04C12N15/867C12N2320/30C12N2740/10041C12N2799/04
Inventor 秦晓峰王子宁纪静芸
Owner FANTASIA BIOPHARMA ZHEJIANG CO LTD