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A method for establishing an immunodeficiency mouse model

An immunodeficient mouse, a technology for establishing a method, applied to other methods of inserting foreign genetic materials, using microinjection, recombinant DNA technology, etc., can solve the problems of long cycle, impure strain background, etc. The effect of shortening the time

Active Publication Date: 2015-08-12
湖南昭泰生物医药有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it takes two years to establish an immunodeficiency mouse strain using this method, which has defects such as a long cycle and impure background of the strain.

Method used

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  • A method for establishing an immunodeficiency mouse model
  • A method for establishing an immunodeficiency mouse model
  • A method for establishing an immunodeficiency mouse model

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Embodiment 1

[0033] Embodiment 1: Use the method of the present invention to make further immune-related gene knockouts to NOD / SCID breed mice, and construct an immunodeficiency mouse model for screening tumor drugs and treating diseases. The method includes the following steps: (1) Using the TEALN gene knockout technique to construct a DNA that disables the function of immune-related genes in mouse cells;

[0034] (2) Purifying the DNA constructed in step (1), and transcribing it into mRNA in vitro using SP6 reverse transcriptase after purification;

[0035] (3) Purify the mRNA obtained in step (2), and freeze it in RNAase-free ultrapure water for use in microinjection;

[0036] (4) Under SPF level feeding conditions, raise pregnant female mice and pseudopregnant female mice;

[0037] (5) Obtain the oviduct from the abdominal cavity of the pregnant female mouse in step (4), wash it with 0.9% normal saline, observe it under a microscope, pick and collect fertilized eggs;

[0038] (6) Injec...

Embodiment 2

[0076] Embodiment 2: Using the method of the present invention to make a further immunodeficiency mouse model for screening tumor drugs and treating diseases on Rag1 gene knockout mice, the method includes the following steps: (1) using TEALN gene knockout, Construction of DNA that disables the function of immune-related genes in mouse cells;

[0077] (2) Purifying the DNA constructed in step (1), and transcribing it into mRNA in vitro using SP6 reverse transcriptase after purification;

[0078] (3) Purify the mRNA obtained in step (2), and freeze it in RNAase-free ultrapure water for use in microinjection;

[0079] (4) Under SPF level feeding conditions, raise pregnant female mice and pseudopregnant female mice;

[0080] (5) Obtain the oviduct from the abdominal cavity of the pregnant female mouse in step (4), wash it with 0.9% normal saline, observe it under a microscope, pick and collect fertilized eggs;

[0081] (6) Inject the mRNA obtained in step (3) into the pronucleu...

Embodiment 3

[0091] Embodiment 3: Using the method of the present invention to make a further immunodeficiency mouse model for screening tumor drugs and treating diseases on Rag2 gene knockout mice, the method includes the following steps: (1) using TEALN gene knockout, Construction of DNA that disables the function of immune-related genes in mouse cells;

[0092] (2) Purifying the DNA constructed in step (1), and transcribing it into mRNA in vitro using SP6 reverse transcriptase after purification;

[0093] (3) Purify the mRNA obtained in step (2), and freeze it in RNAase-free ultrapure water for use in microinjection;

[0094] (4) Under SPF level feeding conditions, raise pregnant female mice and pseudopregnant female mice;

[0095] (5) Obtain the oviduct from the abdominal cavity of the pregnant female mouse in step (4), wash it with 0.9% normal saline, observe it under a microscope, pick and collect fertilized eggs;

[0096] (6) Inject the mRNA obtained in step (3) into the pronucleu...

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Abstract

The invention discloses a method for establishing an immunodeficiency mouse model. The method comprises the following steps of: (1) constructing DNA (deoxyribonucleic acid) which enables functions of an immune related gene in a mouse cell to be lost; (2) purifying mRNA (messenger ribonucleic acid), and cryopreserving in a super-purified water of RNAase free; (4) feeding a pregnant female mouse and a pseudopregnant female mouse under an SPF (specific pathogen free) level feeding condition; (5) taking a fallopian tube from the abdominal cavity of the pregnant female mouse in the step (4), and collecting fertilized ovum; (6) injecting the mRNA obtained in the step (3) into the pronucleus of the fertilized ovum in the step (5) by using a microinjection method; (7) transplanting the fertilized ovum obtained in the step (6) into the body of the pseudopregnant female mouse obtained in the step (4), and enabling the pseudopregnant female mouse to culture the second generation; (8) establishing a stable deficient mouse strain. The method has the advantages that a mouse model which completely does not have immunological rejection to allotransplantation can be obtained for testing.

Description

technical field [0001] The invention relates to a method for establishing an animal model for testing, in particular to a method for establishing an immunodeficiency mouse model. Background technique [0002] The establishment of immune-deficient mouse models is widely used in the screening of tumor drugs, cancer treatment methods and other experiments. The use of immunodeficiency mouse models has a great effect on the study of tumor drugs and the therapeutic mechanism of other diseases, so as to achieve the purpose of preventing and controlling diseases and improving the cure rate. [0003] At present, the breeds commonly used as immunodeficiency mouse models include Rag1 knockout mice, NOD / SCID mice, and Rag2 knockout mice. To measure the degree of immune deficiency in the body, it is mainly to measure the proportion of T, B lymphocytes and NK cells in the body. Rag knockout mice cannot produce mature T and B lymphocytes in vivo, but can produce normal levels of NK cells...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/89A01K67/027
Inventor 李鹏刘志新蒋志武王素娜尹爱兰钟梅贾蓓
Owner 湖南昭泰生物医药有限公司
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