TY  - JOUR
TI  - Measuring Homologous Recombination Frequency in Arabidopsis Seedlings
AU  - Rosa, Marisa  
AU  - Scheid, Ortrun Mittelsten
VL  - 4
IS  - 7
PY  - 2014
DA  - 2014/04/05
SP  - e1094
C1  - Bio-protocol 2014;4:e1094
DO  - 10.21769/BioProtoc.1094
UR  - https://doi.org/10.21769/BioProtoc.1094
AB  - Somatic homologous recombination (SHR) is a major pathway of DNA double-strand break (DSB) repair, in which intact homologous regions are used as a template for the removal of lesions. Its frequency in plants is generally low, as most DSB are removed by non-homologous mechanisms in higher eukaryotes. Nevertheless, SHR frequency has been shown to increase in response to various chemical and physical agents that cause DNA damage and/or alter genome stability (reviewed in March-Díaz and Reyes, 2009). We monitor the frequency of SHR in transgenic Arabidopsis seedlings containing recombination substrates with two truncated but overlapping parts of the β-glucuronidase (GUS) reporter gene (Orel et al., 2003; Schuermann et al., 2005). Upon an SHR event, a functional version of the transgene can be restored (Figure 1A). A histochemical assay applicable to whole plantlets allows the visualization of cells in which the reporter is restored, as the encoded enzyme converts a colorless substrate into a blue compound. This type of reporter has been extensively used to identify gene products required for regulating SHR levels in plants. We analyze plants stimulated for SHR by treatments with DNA damaging agents (bleocin, mitomycin C and UV-C) and compare them to non-treated plants.
KW  - Homologous recombination
KW  - DNA repair
KW  - Genotoxic stress
KW  - Arabidopsis
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False