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Method for identifying goat early embryo sex via PCR

An embryo and goat technology is applied in the field of identifying the sex of goat early embryos, which can solve the problems of difficulty in popularization and use, and achieve the effect of facilitating popularization and use and saving time.

Inactive Publication Date: 2007-06-13
新昌县大东种畜发展有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The technical problem to be solved and the technical task proposed by the present invention are to overcome the defect that existing PCR products need to undergo electrophoresis experiments and then observe and determine gender, which is not easy to be popularized and used in production practice, and provide a non-electrophoretic The Method of Identifying the Gender of Goat Early Embryo by PCR Method

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1: Sex identification of goat 4-cell embryos

[0030] 1. DNA extraction of a small amount of goat 4-cell embryonic cells

[0031] Using the glass needle method, cut 1-2 cells from goat 4-cell embryos, then put the embryonic cell samples into a 200μl centrifuge tube, wash with normal saline and add 10μl sterilized ultrapure water; centrifuge briefly in a centrifuge to separate the cells Place the sample at the bottom of the centrifuge tube and seal the centrifuge tube with a sealing film; immediately place it on ice after boiling at 100°C for 5-10 minutes; after cooling, centrifuge at 12000r / min for 10 minutes, take the supernatant for PCR, and the supernatant contains DNA is the template DNA.

[0032] 2. Specific DNA sequence amplification:

[0033] The total volume of the PCR reaction in the reaction tube is 30 μl, including 1 μl of two specific primers, 200 μM of dNTP, 1U of Taq enzyme, 2.5 mM of MgCl, 3 μl of 10×buffer, 2 μl of template stock solution, 1-1....

Embodiment 2

[0039] Example 2: Sex identification of goat morula stage embryos

[0040] 1. DNA extraction of trace goat morula embryo cells

[0041] Using the glass needle method, cut 2 to 10 cells from goat morula embryos, then put the embryonic cell samples into a 200μl centrifuge tube, wash with normal saline and add 10μl sterilized ultrapure water; centrifuge briefly in a centrifuge, and cell samples Put it at the bottom of the centrifuge tube and seal the centrifuge tube with a sealing film; immediately put it on ice after boiling at 100°C for 5-10 minutes; after cooling, centrifuge at 12000r / min for 10 minutes, take the supernatant for PCR, and the DNA contained in the supernatant is ready as the template DNA.

[0042] 2. Specific DNA amplification

[0043] The total volume of the PCR reaction in the reaction tube is 30 μl, including 1 μl of two specific primers, 200 μM of dNTP, 1 U of Taq enzyme, MgCl 2 2.5mM, 10×buffer 3μl, template stock solution 2μl, EB1~1.5μl, make up 30μl of...

Embodiment 3

[0049] Example 3: Gender Identification of Goat Blastocyst Embryos

[0050] 1. DNA extraction from goat blastocyst-stage embryonic cells

[0051] Use the glass needle method to cut 2-10 cells from the trophoblast cells of goat blastocyst embryos, then put the embryonic cell samples into a 200μl centrifuge tube, wash with normal saline and add 10μl sterilized ultrapure water; briefly centrifuge on the centrifuge , place the fine cell sample at the bottom of the centrifuge tube, seal the centrifuge tube with a parafilm; immediately put it on ice after boiling at 100°C for 5-10 minutes; after cooling, centrifuge at 12,000r / min for 10min, take the supernatant for PCR, supernatant The DNA contained in it is the template DNA.

[0052] 2. Specific DNA amplification

[0053] The total volume of the PCR reaction in the reaction tube is 30 μl, including 1 μl of two specific primers, 200 μM of dNTP, 1U of Taq enzyme, 2.5 mM of MgCl, 3 μl of 10×buffer, 2 μl of template stock solution, 1...

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PUM

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Abstract

The invention relates to the method used PCR to identify the goat early embryo sex. It includes the following steps: sampling from goat early embryo sex to make template DNA; designing primer according to goat SRY gene order; adding the template DNA, primer, DNA polymerase, reaction buffer solution, and dNTP into reaction tube to use as reactant; repeating denaturalization-annealing-stretching three steps to the end of the reaction. Its features are that it adds ethidium bromide into the reaction tube before PCR, directly observes the result after PCR by putting the reaction tube under ultraviolet lamp. It designs a pair of primers according to goat Y chromosome specificity sequence, identifies by non-electrophoresis PCR, simplifies operational programs, and is convenient for popularization and application in production.

Description

technical field [0001] The invention relates to a method for identifying the gender of goat early embryos by using a non-electrophoretic PCR method, which belongs to the technical field of livestock sex control methods. Background technique [0002] Sex control of offspring of livestock has always been one of the subjects of concern, especially for livestock with economic and practical value, such as dairy cows and goats. Therefore, it is of great significance to carry out sex identification on early embryos and accelerate the reproduction of offspring of the desired sex to promote the development of high-efficiency animal husbandry. [0003] PCR (polymerase chain reaction, polymerase chain reaction) amplification technology is a molecular biology method for amplifying the target gene. The enzymatic synthesis reaction relying on DNA polymerase and reaction buffer has the characteristics of fast, specific, sensitive, simple and efficient. PCR is composed of three basic reac...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 徐雪明俞颂东陈阿琴王争光王锡炉张锐
Owner 新昌县大东种畜发展有限公司
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