A construction method and application of plasmids for interfering pig CRTC3 gene expression and adenoviruses

A technology of CRTC3 and construction method, applied in the field of biology and modern agriculture, can solve the problems of unknown regulation mechanism of subcutaneous fat and intramuscular fat difference, and achieve the effect of simple construction method

Inactive Publication Date: 2020-03-06
ZHEJIANG UNIV
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  • Abstract
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  • Application Information

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Problems solved by technology

[0003] However, there is no report on the regulation of CRTC3 on fat deposition in pigs, and whether

Method used

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  • A construction method and application of plasmids for interfering pig CRTC3 gene expression and adenoviruses
  • A construction method and application of plasmids for interfering pig CRTC3 gene expression and adenoviruses
  • A construction method and application of plasmids for interfering pig CRTC3 gene expression and adenoviruses

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Embodiment Construction

[0061] The present invention is further described below in conjunction with specific embodiment, but protection scope of the present invention is not limited thereto:

[0062] (1) Primer design for targeted interference with porcine CRTC3 gene expression

[0063] 1. According to the porcine CRTC3 sequence (Accession: MF964215.1) on NCBI and the carrier information, design 3 pairs of interference primers sh1~3, respectively:

[0064] sh1-F:5-GATCCGGAAGTTCAGCGAGAAGATTGTTCAAGAGACAATCTTCTCGCTGAACTTCCTTTTTTA-3

[0065] sh1-R:5-AGCTTAAAAAAGGAAGTTCAGCGAGAAGATTGTCTCTTGAACAATCTTCTCGCTGAACTTCCG-3

[0066] sh2-F:5-GATCCGGCTAACATCTGCACTCAACATTCAAGAGATGTTGAGTGCAGATGTTAGCCTTTTTTTA-3

[0067] sh2-R:5-AGCTTAAAAAAGGCTAACATCTGCACTCAACATCTCTTGAATGTTGAGTGCAGATGTTAGCCG-3

[0068] sh3-F:5-GATCCGCTCAATAAGACAGCGCTTTCTTCAAGAGAGAAAGCGCTGTCTTATTGAGCTTTTTTA-3

[0069] sh3-R:5-AGCTTAAAAAAGCTCAATAAGACAGCGCTTTCTCTCTTGAAGAAAGCGCTGTCTTATTGAGCG-3

[0070] 2. Three pairs of primers are annealed respectivel...

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Abstract

The invention discloses a construction method of plasmids for interfering pig CRTC3 gene expression. The method comprises the following steps: designing and obtaining three target sequences accordingto the information of a pig CRTC3 gene sequence and a carrier, and designing and synthesizing three pairs of shRNA (1-3) primers of pig CRTC3 according to the target sequences; and connecting annealing products of the three pairs of shRNA (1-3) primers of the pig CRTC3 with pAdM-FH-GFP carriers subjected to BamH I and Hind III enzyme digestion and carrying out conversion to obtain pAdM-FH-GFP-sh(1-3)-CRTC3 recombinant plasmids. The invention further provides an application of the pAdM-FH-GFP-sh1-CRTC3 plasmids and adenoviruses of the pig CRTC3: knocking out the expression of the CRTC3 gene inpig intramuscular fat and subcutaneous fat cells and researching the influence generated by pig glycolipid metabolism and related gene expression and the like.

Description

technical field [0001] The invention relates to a pAdM-FH-GFP-sh1-CRTC3 plasmid that interferes with the expression of porcine CRTC3 (cAMP responsive element binding regulated transcription coactivators) and a construction method and use of the adenovirus, belonging to the application technology in the field of biology and modern agricultural technology. Background technique [0002] The deposition of pig adipose tissue in different parts of the body directly affects the carcass traits and pork quality of pigs. For example, the amount of intramuscular fat deposition affects the meat color, flavor and tenderness of pork; the deposition of subcutaneous fat affects the carcass quality of pork. The fat deposition process in pigs mainly includes the proliferation, differentiation and polyester of preadipocytes, which are regulated by genes related to fat deposition. CRTC3 is a gene highly expressed in adipose tissue. Studies in mice have found that knocking out CRTC3 all over the...

Claims

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Application Information

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IPC IPC(8): C12N15/861C12N15/113C12N15/65C12N7/01
CPCC12N7/00C12N15/113C12N15/65C12N15/86C12N2310/14C12N2710/10021C12N2710/10043
Inventor 单体中刘嘉琪汪以真
Owner ZHEJIANG UNIV
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