Growth differentiation factor 9 (GDF9) is a growth factor secreted by oocytes in growing ovarian follicles, which is essential for growth and differentiation of early ovarian follicles. The polymorphism of GDF9 gene in Small Tail Han sheep, Hu sheep, Dorset sheep and Suffolk sheep was analyzed by PCR-SSCP. The results indicated that there were three genotypes (AA, AB and BB) detected by primer 1. AA genotype was detected in four sheep breeds. AB genotype was detected in Hu sheep, Dorset sheep and Suffolk sheep. BB genotype was only detected in Suffolk sheep. Frequency of A allele was obviously higher than frequency of B allele in four sheep breeds. There were two genotypes (AA and AB) detected by primer 2. AA genotype was detected in four sheep breeds. AB genotype was detected in Hu sheep, Dorset sheep and Suffolk sheep. BB genotype was not detected in four sheep breeds. Frequency of AA genotype was the highest, and frequency of A allele was obviously higher than frequency of B allele in four sheep breeds. The polymorphic fragments amplified by primer 1 were cloned and sequenced. The sequencing results showed that there was one single nucleotide mutation: A-->G at cDNA 152 of GDF9 gene in sheep, and this mutation resulted in an amino acid change: asparagine-->aspartic acid.