The Bombyx mori serves as model organism among the Lepidoptera insects. In the post-genomic era, in order to study gene function, the profiling of mRNA transcription has become a popular research field. Real-time quantitative RT-PCR (qRT-PCR) has become established as the sensitive method for detecting the expression level of low abundance mRNA, and it usually chooses one or several reference genes to standardize the expression level of target gene. Since the changes in amplification of reference gene can reflect the changes of RNA production, quality or cDNA synthesis efficiency. So choosing an appropriate reference gene can reduce the differences between tested samples. Based on the comparison of Standardization of three frequently-used reference genes (GAPDH, Actin-3, 28srRNA), and decide which is the best way to study gene expression level in silkworm, Bombyx mori.