Measurement of geminin activity in Xenopus egg extracts

Methods Mol Biol. 2005:296:263-78. doi: 10.1385/1-59259-857-9:263.

Abstract

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.

MeSH terms

  • Animals
  • Cell Cycle
  • Cell Cycle Proteins / analysis*
  • Cell Cycle Proteins / isolation & purification
  • Cell Cycle Proteins / pharmacology
  • DNA / genetics
  • DNA / isolation & purification
  • DNA Replication / drug effects
  • Female
  • Geminin
  • Male
  • Mitosis
  • Ovum / chemistry*
  • Ovum / cytology
  • Ovum / metabolism
  • Spermatozoa / chemistry
  • Xenopus
  • Xenopus Proteins / analysis*

Substances

  • Cell Cycle Proteins
  • GMNN protein, Xenopus
  • Geminin
  • Xenopus Proteins
  • DNA