Mammalian embryos sex determination method

Abstract

The invention relates to the technical field of sex determination, in particular to a mammalian embryo sex determination method. According to the method, by inserting a fluorescent reporter gene intoa Y chromosome, and by means of fluorescent expression of the fluorescent reporter gene, the embryo sex of a mammal is determined accurately, and the method is simple, efficient, accurate and small indamage; in the process of building a male mammal modal with the Y chromosome containing the fluorescent reporter gene, spacer regions located at Ddx3y and Uty genes are selected, exogenous gene integration is conducted for a target area, and the influence of target site selection on a transgenic animal is lowered; in the gene editing process, by adopting Cas9 protein and single guiding sgRNA designed according to a target area gene sequence, target sites DNA can be cut accurately, the fluorescent reporter gene is inserted between the Ddx3y and Uty genes of the Y chromosome directionally, andthe gene knock-in means is efficient and simple.

Description

【Technical field】 [0001] The invention relates to the technical field of sex identification, in particular to a method for sex identification of mammalian embryos. 【Background technique】 [0002] Sex control technology is a biotechnology that makes adult female animals produce offspring of the desired sex by artificially intervening in the normal reproductive process of animals. This technology is of great significance for improving the production efficiency of limited sex traits in animal husbandry, avoiding the occurrence of sex-linked genetic diseases, and speeding up the breeding process. The discovery of the sex-determining sequence on the Y chromosome is an important breakthrough in the theory of mammalian sex determination, and is of great significance to the development of sex control technology. The current sex control methods mainly focus on two aspects, one is the sperm level and the other is the embryo level. [0003] Separating X and Y sperm and identifying th...

AI Technical Summary

Problems solved by technology

However, the main problem at present is that the separation cost is high, and it is difficult to be widely used in agricultural production.

Other methods for separating X and Y sperm, including electrophoresis, density gradient centrifugation and immunology, have failed to achieve satisfactory results, and the repeatability is poor, so they cannot be used in production

[0004] At present, the most effective methods for gender identification of embryos before implantation in mammals are mainly embryo karyotype analysis and PCR method, but both methods need to remove some cells from the embryo, which will cause a certain degree of damage to the subsequent development of the embryo

In addition, the operation procedure of embryo karyotype analysis is complicated and the cycle is long, which also limits the popularization and application of this method.

Although the PCR method is highly sensitive, it is also prone to contamination, prone to false positives, and has certain limitations in application and promotion.

Although there have been some achievements and developments in sex control techniques, the current limitations of these methods limit their promotion and use in livestock production

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