Study On Sex Identification Of IVF Embryos In Buffalo

As the development of molecular biology technology, the polymerase chain reaction (PCR) has become the main embryo identification method for it's delicacy, speediness and simplification.The application of this method may fully utilize the male or female respective production superiority and enhance the breeding efficiency.In our country, many South provinces have rich water buffalo dairying resources, the development potential is huge, sex identification of water buffalo preimplantation embryo is improtant.But in sex identification procedures, solely amplification of the male unique sex determination gene (Sry) may lead judgement of it as the feminine, when the dual-PCR of double primers are performed, because of single copy of Sry gene and multi- repetitions of euchromosome gene, the Sry gene can not be fully amplified or even amplified.This research adopted double primers and two-step PCR to identify the sex of buffalo by white blood, in this way, to get over the pseudo feminine when single pairs are amplified.when appears, double primers dual PCR amplify when the Sry gene not amplify insufficiency, further enhances the precision of sex identification, optimized sex identification system.Has carried on the discussion about the most suitable embryo template quantity using this optimized sex identification system to the water buffalo preimplantation 16 cells embryos sex identification, and to add the different serum in the embryo culture medium to conduct the research to the water buffalo preimplantation embryo different developmental stage sex ratio influence.Test result as follows.1.Study on water buffalo leucocytes sex identification use double pairs primers dual PCR.sduty on the logical of primer design and the influence double pairs primers dual PCR for sex identification system, to resolve the problem of pseudo feminine caused by use single pair primers amplify and the Sry gene didn't amplify as use double pairs primers dual PCR, optimize the sex identification system.(1) Primer design and its logical examination.The special Y chromosome Sry sequences of male water buffalo and the chromosome 1.715 satellite DNA sequences were attained according to the NCBI gene database, and using the software Primer 5.0 designed two primers S1/S2 and B1/B2, S1/S2 is used to amplify the Sry gene and B1/B2 is used to amplify1.715 satellite DNA sequences.Using this two pairs primers amplify male and female's leucocytes genome, the result showed that the primer S1/S2 can effectively amplify Sry gene which was the male water buffalo's leucocytes had, the fragment length is 350 bp, and no homologous fragment to female leucocytes...