A replication stress-induced synchronization method for Arabidopsis thaliana root meristems

Abstract

Synchronized cell cultures are an indispensable tool for the identification and understanding of key regulators of the cell cycle. Nevertheless, the use of cell cultures has its disadvantages, because it represents an artificial system that does not completely mimic the endogenous conditions that occur in organized meristems. Here, we present a new and easy method for Arabidopsis thaliana root tip synchronization by hydroxyurea treatment. A major advantage of the method is the possibility of investigating available Arabidopsis cell-cycle mutants without the need to generate cell cultures. As a proof of concept, the effects of over-expression of a dominant negative allele of the B-type cyclin-dependent kinase CDKB1;1 gene on cell-cycle progression were tested. The previously observed prolonged G₂ phase was confirmed, but was found to be compensated for by a reduced G₁ phase. Furthermore, altered S-phase kinetics indicated a functional role for CDKB1;1 during the replication process.