Measurement of Geminin Activity in Xenopus Egg Extracts

Geminin is an unstable protein that inhibits DNA replication by interfering with the assembly of prereplication complex at replication origins. Geminin is thought to prevent a second round of replication during late S- or G2 -phase. The protein is destroyed by ubiquitin-dependent proteolysis during mitosis, allowing a new round of replication in the next cell cycle. This chapter describes protocols for measuring the stability of geminin and two of its activities, the inhibition of replication and the inhibition of pre-RC loading.