Cloning Of GDF9 Gene In Goats And Studies On Polymorphisms Analysis In Exons Of GDF9 Gene

Growth differentiation factor 9(GDF9),belonging to TGF-βsuper family, secreted by oocytes in growthing ovarian follicles, plays a central role in the mechanisms of folliculogenesis by regulating granulosa cell proliferation and cyto-differentiation, theca cell formation, and oocyte development, all of which are critical functions for female fertility. The research would provide scientific bases for further genetic reseach on reproduction of goat.One hundred and forty-five Jinning grey goats, sixty Lubei white goats, sixty Laiwu black goats were used. we choose the GDF9 gene as the candidate gene of the breeding trait, then amplified, sequenced, sequence blasted and PCR-SSCP and RFLP tested the exons of different goat populations. The polymorphism of GDF9 gene exons was examined. The results were as follows:1. The primers of GDF9 gene were designed on the base of the sequence of ovine GDF9 gene. Applying PCR and RACE we cloned the gene. The whole gene were 4715bp including two exons and one intron. The nucleotide of GDF9 gene is 98% homologous to sheep.2. To detect DNA variation of sheep GDF9 gene, the sequence of exons has been studied by PCR-SSCP and RFLP, It was showed that there were three mutation sites. After sequencing the E1-2, E2-3 and E2-4, A→C, A→C and G→A transitions were identified at the sites of 1956, 3855 and 4085. The 3855 and 4085 mutations caused amino acid Pro→Gln and Ile→Val respectively.3. Polymorphisms of GDF9 gene were as follows. For primer 1-2, three genotypes (AA, AC and CC) were detected, the polymorphism information content (PIC) showed that the Lubei white goat population was lower degree polymorphism, Laiwu black goat and Jining green goat populations were moderate polymorphism. For primer 2-3, three genotypes (AA, AC and CC) were detected, the polymorphism information content (PIC) showed that the Lubei white goat and Laiwu black goat populations were lower degree polymorphism, and Jining green goat population was moderate polymorphism. For primer 2-4, three genotypes (AA, AT and TT) were detected, the polymorphism information content (PIC) showed that the Lubei white goat and Laiwu black goat populations were moderate polymorphism and Jining green goat population was lower degree polymorphism.