Abstract: Suppression subtractive hybridization technology and real-time quantitative PCR were used to isolate differential expressed genes in peak lactation stage.Results showed that peak to early subtraction library was successfully built with cDNA from mammary gland of Xinong Saanen goat at the two lactation stages, and the subtraction efficiency was approximately 2⁵ fold indicated by GAPDH; PCR analysis of the total 78 positive clones demonstrated that the cDNA inserts range largely from 150 bp to 1 000 bp; 25 cDNA sequences were identified from 30 clones sequenced which represent 18 genes from the peak to early SSH library; Moreover four genes namely serum amyloid A protein 3 (SAA3), ATP-binding cassette subfamily G member 2 (ABCG2), heart fatty acid-binding protein (H-FABP) and xanthine dehydrogenase (XDH) were chosen to do real-time quantitative PCR to confirm the expression differentiation, and found they were 17.0, 7.7, 16.3 and 1.7 fold higher in mammary gland of peak lactation than that of early lactation, respectively. Conclusion: the constructed subtraction library could be used to identify different expression genes in peak lactation stage, and the 4 genes verified by real-time quantitative PCR might become candidate genes which had function in regulating the change of milk yield and composition.