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Non-alcoholic fatty liver mouse model and construction method thereof

A mouse model, non-alcoholic technology, applied in the field of modeling technology of medical animal models, can solve the problems of low modeling success rate, long modeling time, limited models, etc., achieve short modeling time and reduce fat decomposition , the effect of increased fat synthesis

Pending Publication Date: 2021-02-12
DALIAN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to provide a mouse model of non-alcoholic fatty liver and its construction method to solve the problem that the current animal models of non-alcoholic fatty liver used in scientific research are very limited, and there is no non-alcoholic fatty liver with stable heredity in the market. Liver model mice all need to be cultivated and modified the day after tomorrow. The modeling time is long, the success rate of modeling is low, and the mortality rate is high.

Method used

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  • Non-alcoholic fatty liver mouse model and construction method thereof
  • Non-alcoholic fatty liver mouse model and construction method thereof
  • Non-alcoholic fatty liver mouse model and construction method thereof

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Embodiment 1

[0029] The present invention provides such Figure 1-3 A non-alcoholic fatty liver mouse model and its construction method are shown, and the specific construction steps are:

[0030] Step 1: transfect mouse embryonic stem cells, use Pfu Turbo DNA polymerase to amplify and clone into the pCR Blunt II-TOPO vector containing the 4th and 5th exon fragment (size is 10.8kb) of ghr gene, described The pCRBlunt II-TOPO vector was purchased from Invitrogen, USA, and an 8kb fragment was excised from it with BamHI and XbaI, and then connected to the PL253 vector containing a thymidine kinase cassette (Thymidine kinase cassette) using T4 ligase. The PL452 and PL451 vectors containing the neomycin (neo) cassette were recombined and exchanged for this subcloned genomic region, and 2 Loxp sites and the Frt-neo-Frt-LoxP cassette were inserted to obtain a conditional targeting vector, which was restricted by Not I The conditional targeting vector was linearized by endonuclease digestion, and...

Embodiment 2

[0037] The present invention provides such Figure 1-6 A non-alcoholic fatty liver mouse model and its construction method are shown, and the specific construction steps are:

[0038] Step 1: transfect mouse embryonic stem cells, use Pfu Turbo DNA polymerase to amplify and clone into the pCR Blunt II-TOPO vector containing the 4th and 5th exon fragment (size is 10.8kb) of ghr gene, described The pCRBlunt II-TOPO vector was purchased from Invitrogen, USA, and an 8kb fragment was excised from it with BamHI and XbaI, and then connected to the PL253 vector containing a thymidine kinase cassette (Thymidine kinase cassette) using T4 ligase. The PL452 and PL451 vectors containing the neomycin (neo) cassette were recombined and exchanged for this subcloned genomic region, and 2 Loxp sites and the Frt-neo-Frt-LoxP cassette were inserted to obtain a conditional targeting vector, which was restricted by Not I The conditional targeting vector was linearized by endonuclease digestion, and...

Embodiment 3

[0045] The present invention provides such Figure 1-8 A non-alcoholic fatty liver mouse model and its construction method are shown, and the specific construction steps are:

[0046] Step 1: transfect mouse embryonic stem cells, use Pfu Turbo DNA polymerase to amplify and clone into the pCR Blunt II-TOPO vector containing the 4th and 5th exon fragment (size is 10.8kb) of ghr gene, described The pCRBlunt II-TOPO vector was purchased from Invitrogen, USA, and an 8kb fragment was excised from it with BamHI and XbaI, and then connected to the PL253 vector containing a thymidine kinase cassette (Thymidine kinase cassette) using T4 ligase, and then The subcloned genomic region was recombined and exchanged with PL452 and PL451 vectors containing a neomycin (neo) cassette, and two Loxp sites and a Frt-neo-Frt-LoxP cassette were inserted to obtain a conditional targeting vector, which was passed Not I Linearize the conditional targeting vector by restriction endonuclease digestion an...

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Abstract

The invention discloses a non-alcoholic fatty liver mouse model and a construction method thereof. The construction method includes the specific construction step that 1, mouse embryonic stem cells are transfected, and a fourth exon segment and a fifth exon segment containing ghr genes are amplified through Pfu Turbo DNA polymerase and cloned into a pCR Blunt II-TOPO vector. When a mouse modeled through the method is 8 weeks old, the liver volume is remarkably increased, H&E staining and oil red staining results show fatty liver lesion, liver cell fat synthesis is remarkably increased, lipolysis is remarkably reduced, and the mouse model built through the method has the advantages of being stable in heredity, similar to human conditions in clinical symptoms, short in modeling time, high in efficiency and low in death rate. The model is capable of being formed without diet induction, and relatively wide in application and popularization prospect, and is an ideal non-alcoholic fattyliver animal model for drug screening and pharmacological research.

Description

technical field [0001] The invention relates to the technical field of modeling techniques for medical animal models, in particular to a non-alcoholic fatty liver mouse model and a construction method thereof. Background technique [0002] Non-alcoholic fatty liver disease (NAFLD) is a type of chronic liver disease characterized by diffuse hepatocyte hypertrophy and vesicular hepatic steatosis. In recent years, the number of NAFLD patients has been increasing year by year. It has seriously threatened human health. The "Guidelines for the Prevention and Treatment of Fatty Liver in China (Popular Science Edition)" (2018 Edition) shows that the prevalence of non-alcoholic fatty liver in China is as high as 25%, and 15% of them develop into non-alcoholic steatohepatitis , a small number of patients develop liver cirrhosis and liver cancer. Non-alcoholic fatty liver disease is more complicated. If it is not controlled in time, it will lead to inflammation and liver fibrosis. Pat...

Claims

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Application Information

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IPC IPC(8): A01K67/027C12N15/85
CPCA01K67/0276C07K14/47C12N15/85A01K2217/075A01K2227/105A01K2267/03
Inventor 吴英杰孙杰姜如娇冉丽媛郭美华于东
Owner DALIAN MEDICAL UNIVERSITY
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