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Method for improving pig meat quality

A technology for muscle quality and muscle tissue, applied in the field of improving pig muscle quality, can solve the problems of verifying the individual biological function of transgenic pigs, and methods that have not yet been seen in pig muscle quality, so as to increase the intramuscular fat content of pigs and improve muscle mass. quality effect

Inactive Publication Date: 2015-11-11
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In 2013, Zhu Yun et al. found that "different genotypes of the BsrI site of the PPARγ2 gene have significant differences in fat traits such as intramuscular fat content and shear force, but there is no significant difference in marbling"
However, the exogenous gene used in the construction of the pN1-MCK-PPARγ2 vector is the PPARγ2 gene from pigs, and the biological function has not been verified in transgenic pigs.
[0010] So far, there has been no report on the method of improving pig muscle quality by transgenic PPARγ2

Method used

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  • Method for improving pig meat quality
  • Method for improving pig meat quality
  • Method for improving pig meat quality

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1: Establishment of porcine embryonic fibroblast cell line

[0042] The 30-day embryonic uterus of large white pigs were taken, stored in an incubator, and brought back to the laboratory within 2 hours. Separate the fetus from the uterus and put it in a phosphate buffer solution containing 1× double antibody, that is, PBS (the formula is 8.0gNaCl, 0.2gKCl, 2.9gNa 2 HPO 4 .12H 2 O, 0.2gKH 2 PO 4 , double-distilled water 1000ml: the double-antibody is 10,000units / ml of penicillin and 10,000ug / ml of streptomycin) in a beaker, and then transferred to the ultra-clean bench for operation, and the fetal head, limbs, and viscera were cut off with ophthalmology, and placed in Store in a 2ml EP tube at -80°C for later use, wash three times with the PBS containing the double antibody, try to wash until there is no blood, and then wash once with dimethyl sulfoxide (DMEM) solution to remove the influence of the double antibody; transfer to a new culture Cut the remainin...

Embodiment 2

[0044] Example 2: Establishment and detection of transgenic cell lines

[0045] Take out a bottle of cultured 30-day-old pig fetal fibroblasts, digest, rinse and centrifuge, add 350 μl of serum-free medium (invitrogen, purchased from Wuhan Dafeng Biotechnology Co., Ltd.), and then add 5 μg of linearized pN1- MCK-PPARγ vector (for the construction of the vector, see figure 2 , the vector construction method is to remove the GFP gene from the eukaryotic expression vector pEGFP-N1 as the framework, and then use the gene capture technology from the BAC of the pig MCK gene to capture the 7Kb promoter MCK sequence and construct the vector pN1-MCK. Double digestion with SalI and NotI to obtain a large fragment pN1-MCK. The artificially synthesized PPARγ2 cDNA full-length plasmid (pMD-18-PPARγ2) was used as material, and the PPARγ2 gene fragment was obtained by performing double enzyme digestion with SalI and NotI respectively. The large fragment pN1L-MCK and PPARγ2 cDNA obtained ...

Embodiment 3

[0047] Embodiment 3: the preparation of transgenic pig

[0048] Take the ovary of a sow that has just been slaughtered, put it into a thermos cup filled with physiological saline containing penicillin / streptomycin at 38°C, and transport it back to the laboratory within 2 hours. The follicular fluid was extracted, and the cumulus oocyte complex with regular shape, closely surrounded by more than 3 layers of cumulus cells and dense and uniform cytoplasm was selected under a stereoscope, and placed in the in vitro maturation culture medium (the formula was MAT mother solution 8.5 ml, 500ul cysteine ​​(0.0070g dissolved in 5ml MAT mother solution), 1mlpFF, 10ulEGF (0.05mg / ml, dissolved in mature solution), 10ulLH (0.5mg / ml, dissolved in DPBS), 10ulFSH (0.5mg / ml, dissolved in DPBS) .) After rinsing 3 times, transfer to mineral oil to cover, at least in CO 2 The in vitro maturation medium in the incubator that has been balanced for 3-4 hours is cultured for 42 hours, and the aggreg...

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Abstract

The invention belongs to the technical field of animal genetic engineering and particularly relates to a method for improving pig meat quality. The method is characterized in that a peroxisome proliferator-activated receptor (PPAR) gamma gene serves as an important candidate gene for improving the meat quality, a fibroblast cell line of a 30-day-old fetus of a large white pig is established, a SwaI linearized pN1-MCK-PPAR gamma2 expression vector is shifted to the 30-day-old fetus fibroblast cell line of the large white pig through an electrotransfection method, and a PPAR gamma transgenic pig is prepared in a method of somatic nucleus transplantation. The influence of muscle tissue overexpression PPAR gamma genes on meat traits such as intramuscular fat deposition is verified in a transgenic pig, the contradiction of simultaneously selecting meat quality and meat quantity in conventional animal breeding is overcome, and the novel method is provided for cultivating lean meat pigs with good meat quality.

Description

technical field [0001] The invention belongs to the technical field of animal genetic engineering, and in particular relates to a method for improving pig muscle quality. The present invention constructs a pig skeletal muscle-specific promoter MCK and peroxisome proliferator-activated receptor gamma (PPARγ) combined with a p-N1 carrier to construct a muscle-specific induction PPARγ overexpression system, which is introduced by electroporation Pig embryonic fibroblasts, and transgenic pigs were prepared by somatic cell nuclear transfer technology, and their effects on intramuscular fat and pork quality were verified at the individual level of transgenic pigs, laying a technical foundation for breeding new breeds of lean pigs with good muscle quality. Background technique [0002] Transgenic animals (Transgenic animals) refer to the use of bioengineering methods to introduce and stably integrate the constructed exogenous target gene fragments into the recipient chromosome geno...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/873A01K67/027
Inventor 熊远著周颖左波徐德全任竹青夏晓亮马志远
Owner HUAZHONG AGRI UNIV
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