Application of abcg4 gene in constructing animal model with two-way change in obesity degree

A technology of genes and uses, applied in the field of Abcg4 gene in the construction of animal models with bidirectional changes in the degree of obesity

Active Publication Date: 2021-12-24
JIANGSU UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the obese animal models used for a long time have no difference in the degree of obesity between different genders, and so far there are no related reports on animal models with bidirectional changes in the degree of obesity between different sexes

Method used

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  • Application of abcg4 gene in constructing animal model with two-way change in obesity degree
  • Application of abcg4 gene in constructing animal model with two-way change in obesity degree
  • Application of abcg4 gene in constructing animal model with two-way change in obesity degree

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] see figure 1 , the method for knocking out mouse Abcg4 gene of the present embodiment, comprises the steps:

[0042] Using the wild-type mouse genome as a template, the primer pair I was used to amplify the fragment comprising the 6th exon to part of the 7th exon of the Abcg4 gene, denoted as fragment I;

[0043] The nucleotide sequence of the primer pair I is:

[0044] FI:5'-TGGTGGATGGTGCAGATGAC-3' (as shown in SEQ ID NO.1);

[0045] RI: 5'-AAATCGGGGTGGTGCTTAGG-3' (as shown in SEQ ID NO.2);

[0046] Amplify the fragment comprising the 9th-14th exon of the Abcg4 gene with primer pair II, denoted as fragment II;

[0047] The nucleotide sequence of the primer pair II is:

[0048] FII:5'-AACTGACGAAGAAGCCGGAG-3' (as shown in SEQ ID NO.3);

[0049] RII: 5'-ATCAAAGAAGGACTGCCCCG-3' (as shown in SEQ ID NO.4);

[0050] Amplify the fragment comprising the 9th-14th exon of the Abcg4 gene with primer pair II, denoted as fragment II;

[0051] S2. Insert fragment I into the eG...

Embodiment 2

[0059] Genotype identification of Abcg4 gene-deficient mice

[0060] Confirm the correctness of homologous recombination with Southern hybridization, including the following steps:

[0061] The genomic DNA of the Abcg4 gene-deficient mouse obtained in Example 1 and the genomic DNA of the wild-type mouse were first digested with BgII enzyme, and probes were made for Southern hybridization detection.

[0062] The preparation method of the probe is as follows: PCR amplification of genome DNA is first performed with primers int2F-2 and int5R. The sequence information of int2F-2 and int5R is as follows:

[0063] int2F: 5'-CAGTACACAGGTTAAGCTAGGCAG-3' (as shown in SEQ ID NO.5)

[0064] int5R: 5'-TCCATACACAGGCCTCGCCTAAGC-3' (as shown in SEQ ID NO.6)

[0065] The PCR reaction system is: DNA template 10ng, primers 2ug, Taq enzyme 1unit, 1×PCR buffer, and the total reaction volume is 20ul.

[0066] PCR reaction conditions: 94°C, 5 minutes; cycle: 94°C, 30 seconds, 60°C, 45 seconds, 7...

Embodiment 3

[0069] Characterization of Abcg4 gene-deficient mice using indicators of obesity

[0070] The following results were compared with Abcg4 gene-deficient mice and wild-type mice born in the same littermate at 4 weeks after birth.

[0071] (1) Analysis of triglyceride content

[0072] Analysis process and results: using the triglyceride analysis kit from Boehringer Mannheim and operating according to its instructions, the results are as follows image 3 shown.

[0073] (2) Analysis of paragonadal fat pad

[0074] Analysis process and results: mice were anesthetized, weighed and recorded body weight, dissected mice, took out paragonadal fat pad, weighed and recorded, the results were as follows: Figure 4 shown. The ratio of the fat pad weight to the body weight of the mouse is the fat content rate (in percentage), and the results are as follows Figure 5 shown.

[0075] (3) Pancreas weight analysis

[0076] Analysis process and results: the mice were dissected to take out ...

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Abstract

The invention belongs to the field of animal genetic engineering and genetic modification, and in particular relates to the application of Abcg4 gene in constructing an animal model with two-way changes in obesity degree. The use specifically refers to constructing an animal model by knocking out the Abcg4 gene in an animal, or inhibiting the expression of the Abcg4 gene. The degree of obesity in the animal model is dependent on sex, as compared to wild-type littermates of the same sex, females become obese and blood triglyceride levels decrease, while males become lean and blood triglyceride levels increase. The model of the invention can be used to study obesity and related diseases, and can be used to screen out more effective and targeted drugs for treating obesity or treating elevated plasma triglycerides.

Description

technical field [0001] The invention belongs to the field of animal genetic engineering and genetic modification, and specifically relates to the application of Abcg4 gene in constructing an animal model with two-way changes in obesity degree. Background technique [0002] Sugars and lipids are essential nutrients for the human body. They not only provide energy for cells, but also are the basic structural components of cells. Glucose and lipid metabolism can be precisely regulated in healthy individuals, and abnormal glucose and lipid metabolism can cause many diseases including obesity, fatty liver, diabetes, atherosclerosis, cardiovascular and cerebrovascular diseases, and even cancer. Therefore, the homeostasis of glucose and lipid metabolism plays an important role in the health of the body. [0003] Obesity is a complex disease caused by the excessive accumulation of body fat, and the obesity gene refers to that the protein encoded by the gene is a component of an app...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85A01K67/027
CPCC12N15/8509A01K67/0276C07K14/70596C12N2800/107A01K2267/0362A01K2227/105A01K2217/054C07K14/705
Inventor 吴金美
Owner JIANGSU UNIV OF SCI & TECH
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