Method for producing transgenic buffalo embryos by applying intracytoplasmic sperm injection (ICSI) mediation

A transgenic, buffalo technology, applied in the field of genetically engineered transgenic animals, can solve the problems of low milk production and reproduction rate, restricting the development of buffalo milk industry, etc.

Inactive Publication Date: 2011-02-16
GUANGXI UNIV
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  • Summary
  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, the milk yield and reproduction rate of buffalo breeds in my country are low, which seriously restricts the development of buffalo milk industry

Method used

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  • Method for producing transgenic buffalo embryos by applying intracytoplasmic sperm injection (ICSI) mediation
  • Method for producing transgenic buffalo embryos by applying intracytoplasmic sperm injection (ICSI) mediation
  • Method for producing transgenic buffalo embryos by applying intracytoplasmic sperm injection (ICSI) mediation

Examples

Experimental program
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Effect test

Embodiment 1

[0022] From 2008 to 2009, among a batch of 102 injected eggs, 63 split (61.8%), 38 blastomere EGFP-expressed embryos (60.3%), and 17 blastocysts were obtained, of which 11 blastocysts EGFP was expressed, and the transgenic positive rate of blastocysts was as high as 64.7%. One of the transgenic embryos was cultured in vitro for a long time, and the expression of EGFP could still be seen at 27 days, which proved that the application of this method can effectively produce transgenic buffalo embryos .

Embodiment 2

[0024] Between 2008 and 2009, we injected 95 oocytes, and after activation, 86 of them split (90.5%), 36 of which expressed EGFP (41.9%), and 16 blastocysts were obtained after 7 days of in vitro culture. 10 of them expressed EGFP (62.5%). This example proves that the method of the present invention for producing transgenic buffalo embryos through ICST-mediated method is feasible.

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Abstract

The invention relates to a method for producing transgenic buffalo embryos by applying intracytoplasmic sperm injection (ICSI) mediation. The method comprises the following steps of: 1, incubating buffalo sperms and transgenic deoxyribonucleic acid (DNA) together; 2, performing intracytoplasmic microinjection on the buffalo sperms; and 3, cultivating and identifying the transgenic buffalo embryos. By the method of the invention, the transgenic buffalo embryos are obtained successfully. In 102 injection spawns of the transgenic buffalo embryos, 61.8 percent of the injection spawns are subjected to cleavage, 17 blastospheres are obtained in embryos expressed by blastomere serving as enhanced green fluorescent protein (EGFP), 11 blastospheres express the EGFP, and the transgenic positive rate of the blastospheres is up to 64.7 percent; and after one of the transgenic embryos is cultured in vitro for a long time, the expression of the EGFP can be still seen during 2 to 7 days, so the method makes the important progress for producing transgenic buffalos further.

Description

technical field [0001] The invention belongs to the genetically engineered transgenic animal technology, in particular to a method for producing transgenic buffalo embryos mediated by ICSI. Background technique [0002] Buffalo is an important livestock with great development prospects in southern my country, because of its strong adaptability, high temperature and high humidity resistance, strong disease resistance, resistance to rough feeding, easy feeding, long service life and high nutritional value of dairy products, it has become more and more popular. people's attention and attention. However, at present, the milk yield and reproductive rate of buffalo varieties in my country are low, which seriously restricts the development of buffalo milk industry. ICSI-mediated gene transfer (ICSI-Mediated Gene Transfer, ICSI-Tr) technology, through the preparation of high-purity transgenic vectors and sperm incubation, and then intracytoplasmic sperm microinjection fertilization,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/89C12N15/877
Inventor 石德顺陈自红刘庆友
Owner GUANGXI UNIV
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