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Non-human mammal model with input UCP1-1cuiferase (uncoupling protein 1-1cuiferase) gene as well as construction method and application thereof

A non-human mammal and uncoupling protein technology, applied in the field of non-human mammal model and its construction, can solve the problems of time-consuming, labor-intensive, labor-intensive and expensive, and achieve simple results

Active Publication Date: 2014-07-30
GUANGZHOU INST OF BIOMEDICINE & HEALTH CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, no matter whether real-time quantitative RT-PCR or western blot is used for drug screening, it is time-consuming, labor-intensive and expensive.

Method used

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  • Non-human mammal model with input UCP1-1cuiferase (uncoupling protein 1-1cuiferase) gene as well as construction method and application thereof
  • Non-human mammal model with input UCP1-1cuiferase (uncoupling protein 1-1cuiferase) gene as well as construction method and application thereof
  • Non-human mammal model with input UCP1-1cuiferase (uncoupling protein 1-1cuiferase) gene as well as construction method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1 knocks in the construction method of the mouse model that has uncoupling protein 1-luciferase

[0029] Such as figure 1 As shown, it is a construction strategy diagram of the mouse model knocked in with UCP1-lcuiferase in this embodiment, including the following steps:

[0030] 1. Remove the stop codon of UCP1

[0031] UCP1 has 6 exons, and the last exon of UCP1, that is, the stop codon after the 6th exon, is removed;

[0032] 2. Link luciferase gene

[0033] The tandem peptide 2A (SEQ ID NO: 1, whose purpose is to allow the luciferase gene to be translated into a protein together with UCP1, but not fused with the UCP1 protein, so it will not affect the activity of the UCP1 protein) is removed in step 1 The back of the last exon of UCP1 of the stop codon is connected into the luciferase gene; the sequence after the connection of the luciferase gene is shown in SEQ ID NO:2;

[0034] 3. Obtain the mouse model

[0035] A mouse model with UCP1-luciferase k...

Embodiment 4 Embodiment 1

[0066] Example 4 In vivo imaging detection of knock-in uncoupling protein 1-luciferase mouse model of Example 1

[0067] Include the following steps:

[0068] 1) Prepare D-luciferin Firefly stock solution (15mg / ml), that is, fully dissolve luciferin in DPBS, and then filter with a 0.22 micron filter membrane. Store at -20°C.

[0069] 2) Use Avertin (first weigh 1.25g of tribromoethanol, add 2.5ml of tert-amyl alcohol, then add 97.5ml of ultrapure water, and stir overnight in the dark (at room temperature). Filter with a 0.22μm filter the next day, and pack 4 ℃ protected from light) anesthetized a mouse (weight 24.7 g) knocked in with uncoupling protein 1-luciferase;

[0070] 3) Inject 247 microliters of fluorescein stock solution 10 to 15 minutes before imaging (a mouse is injected at a dose of 10 microliters per gram);

[0071] 4) Perform in vivo imaging on a fluorescence imager.

[0072] In vivo imaging such as Figure 4 shown, from Figure 4 It can be seen that the br...

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Abstract

The invention discloses a non-human mammal model with an input UCP1-1cuiferase (uncoupling protein 1-1cuiferase) gene as well as a construction method and the application of the non-human mammal model. The constructed non-human mammal model with the input UCP1-1cuiferase gene can be taken as a screening platform of white fat cell brownness drugs, and the simple, rapid and large-scale screening on drugs for regulating and controlling the expression of the UCP1 can be realized; on the premise that non-human mammal is not hurt, the expression and distribution of the UCP1 can be directly subjected to bio-assay by utilizing a fluorescent imager.

Description

technical field [0001] The invention belongs to the field of biotechnology, and more specifically, the invention relates to a non-human mammal model knocked in with an uncoupling protein 1-luciferase gene, its construction method and application. Background technique [0002] It has long been thought that the main function of fat cells is to store energy. However, with the in-depth study of fat cells, other functions of fat cells have gradually emerged. Studies have shown that adipocytes play an important role in metabolic diseases such as immune response, hypertension, obesity, and insulin resistance. [0003] Initially, fat cells were divided into two types: white fat cells and brown fat cells. However, with the discovery of beige (brown-in-white, brite) cells in the inguinal fat of rodents, brown adipocytes can be subdivided into two types: classic brown adipocytes and brite / beige adipocytes [0004] White adipocytes contain a large, round lipid droplet with a flattene...

Claims

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Application Information

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IPC IPC(8): A01K67/027C12N15/85
CPCA01K67/027C12N15/85
Inventor 吴东海聂涛毛刘峰李快唐小凤惠晓艳刘彭涛徐爱民
Owner GUANGZHOU INST OF BIOMEDICINE & HEALTH CHINESE ACAD OF SCI
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