Non-radioisotopic quantitative RT-PCR to detect changes in mRNA levels during early mouse embryo development.

We developed a non-radioisotopic quantitative RT-PCR method with high sensitivity and reproducibility. The results of this RT-PCR were in agreement with those of the Northern blot analysis. We measured the mRNA levels of beta-actin, transferrin receptor, and two cell cycle-related genes, cyclin B and cdc25, in early mouse embryos by the RT-PCR. In late two-cell stage embryos, beta-actin, transferrin receptor and cyclin B mRNA levels were 10-20% of those in MII stage oocytes. In contrast, the cdc25 mRNA levels were not different between these stages. When we cultured mouse embryos, the presence of an RNA polymerase inhibitor, alpha-amanitin, in the medium did not affect the mRNA levels at the two-cell stage, indicating that most of the detected mRNAs in two-cell embryos were maternally derived. These results suggest that the rate of mRNA degradation is different between cyclin B and cdc25 during early embryogenesis.