Research Of Candidate Genes Related To Prolific Character On The Goats

China is a big country on goats' feeding. But the reproductive rate of goats are commonly low. Most of them can only born 1～2 lambs in each pregnancy. This remarkably obstacled the production and development of commercial goats. The best way to solve this problem was to enhance the selective breeding, carry out the project of good breed, perfect the reproduction system of good breed on the base of good pure breed. Searching the molecular genetic marker or main gene of high fecundity in goat and applying them to the MAS was the powerful way to carry out the project of good breed fast. So we designed this experiment using the candidate gene method, choosed four candidate genes related to reproduction, such as two prolific main genes in sheep FecX~B and FecG~H which were found by Hanrahan (2004), the NO.10 exon of FSHR gene and the 5' regulative region of FSHβgene, let the high fecundity breed of DaZu black goat, the middle fecundity breed of JinTang black goat, and the low fecundity breeds of ChuanDong white goat, NanJiang yellow goat and BanJiao goat to be our research objects. We Planed to search the polymorphic distribution of the four gene fragments among the five kinds of goats expecting to linked them with the reproductive traits of DaZu black goat for analysis, in order to be helpful to the reservation, breeding and improving of DaZu black goat and other goats' breeds, and to provide some useful molecular information for southern goats' high fecundity.Our experiment had two parts, one part was to use PCR-RFLP method to search the Cambridge and Belclar sheep's high fertility genes FecX~B and FecG~H on five southern goats' breeds such an DaZu black goat et al.Another part was to use PCR-SSCP method to detect the polymorphism of the NO. 10 exon of FSHR gene and the 5' regulative region of FSHP gene on the five goats' breeds.The results were as follows:(1) Using restrictive enzyme Dde I to digest the PCR production of FecG (G8 mutation) and FecX (B4mutation) gene fragments. All the samples were totally been digested. All of the five kinds of goats were wild purity ,there did not exist any mutation site in these five kinds of goats.(2) Using the PCR-SSCP method for the PCR production of NO. 10 exon of FSHR gene, we obtained the coincident result in five kinds of goats, they all showed four band on the glue. There was no diversity between different goats.(3) Aanalyzed the 5'regulative region of FSHp gene using the PCR-SSCP method, we found three different phenotype, they were there, four and five bands phenotype. We judged them as AA,BB, AB. genotype respectively. The BB genotype is the predominant type, then the AB genotype and the AA genotypewas the least. DaZu black goat had two genorypes(BB,AB);Jin Tang black goat had two genotypes(AA,BB,AB);ChuanDong white goat had only one genotype(BB); Banjiao goat had two genotypes(BB,AB);NanJiang yellow goat had two genorypes(BB,AB).These genotypes were in Hardy-Weinberg balance state in all the five goats' breeds. The test of homozygosity, heterozygosity, effective number of alleles, polymorphism information content (PIC)revealed that JinTang black goat had the highest score in every index ,ChuanDong white gaot had the lowest. The mutative extent from high to low was JinTang black goat＞BanJao gaot＞NanJiang yellow goat＞DaZu black goat＞ChuanDong white goat. Only the JinTang black goat reached the middle level of polymorphism information content at 0.25＜PIC＜0. 5, the other breeds had very low PIC, some even had not any PIC at all.(4) According to sequencing results about the NO.10 exon of FSHR gene and the 5'regulative region of FSH(3 gene we found that the NO. 10 exon of FSHR gene are 100% similar to the already known goat sequence in Genebank.The similarity with sheep,cattle,swine and human were 97%,96%,93%,93% respectly. The 5'regulative region of FSHp gene had the similarity with sheep, cattle.swine and human at the percentage of 98%,95%,93% and 89% respectly. Further more we found there was a C-G mutation site at 102bp.The online motif analysis of FSHβgene 5'regulative region showed that there were 8 likely motifs in 13 sites, but the mutation site did not locate in the motif sequenc.With the combination of our research results ,we can drew the conclusion as follows:(1) In five goats breeds such as DaZu black goat, JinTang black goat,BanJiao gaot,NanJiang yellow goat, ChuanDong white goat there were no FecX~B and FecG~H gene been found, and did not exist polymorphism on NO. 10 exon of FSHR gene . All these indicated that the three gene fragments we shoosed did not have anything to do with the high fecundity of the goats we researched.(2) The experiments revealed that there was polymorphism on FSHp gene 5'regulative region.There were three genotypes,these genotypes were in Hardy-Weinberg balance state in all the five goats' breeds,they did not be influenced by the artificial breeding.The polymorphism information content(PIC) from high to low was JinTang black goat＞BanJiao gaot＞NanJiang yellow goat＞DaZu black goat＞ChuanDong white goat.This rank did not have any notable linear relation with the reported reproductive traits of these goats' breeds.It could primarily eliminate the influence of FSHβgene 5'regulative region on goats' fertility.(3) The sequencing results of NO. 10 exon of FSHR gene and the 5' regulative region of FSHβgene further firstly revealed a C-T mutation site in FSHp gene 5' regulative region ,but the mutation site did not exist any motif.This meaned that the mutation might not influence the transcription of FSHβgene.According to the Blast results, the NO. 10 exon of FSHR gene had the similarity with sheep,cattle,swine and human at the rate of 97%,96%,93%,93% respectly. The 5'regulative region of FSHβgene had the similarity with sheep, cattle,swine and human at the rate of 98%,95%,93% and 89% respectly.They all showed a decline tend.This further proved that goat had a increasing genetic distance with sheep, cattle,swine and human.