Abstract
Background: The plateau-type Tibetan sheep is a vital livestock resource in China’s alpine regions and its reproductive performance directly impacts population expansion and economic value. The GDF9 gene is a key regulator of mammalian reproductive traits, with its SNPs closely associated with litter size. This study aimed to establish a PCR-SSCP method for SNP detection in the GDF9 gene of plateau-type Tibetan sheep, providing an efficient and economical tool for genetic breeding. Methods: Specific primers were designed for the GDF9 exon region to amplify target DNA fragments. PCR products were analyzed using SSCP with non-denaturing polyacrylamide gel electrophoresis to screen for SNPs based on band mobility shifts and genotype and allele frequencies were calculated. Result: Stable and clear PCR-SSCP profiles were obtained under the optimized PCR and SSCP conditions. A c.1111G greater than A SNP in exon 2 of the GDF9 gene was identified and verified by sequencing. Two genotypes, AA and AB, were detected, with genotype frequencies of 0.40 and 0.60, respectively, whereas the BB genotype was not observed. The mean litter size of AB-type ewes (1.85±0.18) was significantly higher than that of AA-type ewes (1.52±0.15) (P = 0.035). These results indicate that the established PCR-SSCP method can effectively detect the GDF9 SNP in plateau-type Tibetan sheep and provide a basis for further validation of its association with reproductive traits.