Arabidopsis CAF-1 depletion / gamma-irradiation

CAF-1 is involved in nucleosome assembly following DNA replication and nucleotide excision repair. In Arabidopsis thaliana, the 3 CAF-1 subunits p150, p60 and p48 are encoded by FAS1, FAS2 and, most likely, MSI1, respectively.fas mutants exhibited increased levels of DNA double-strand breaks, a G2 phase retardation, and elevated levels of mRNAs coding for proteins involved in homologous recombinational repair. The same genes that were transcriptionally up-regulated in the fas mutants were also found to have a higher steady state level of transcription in wild-type Arabidopsis plants exposed to gamma-irradiation. Keywords: stress response, genetic modification Total RNA prepared from 4-week-old seedlings (without the roots i.e. true leaves with cotyledon and hypocotyls) using an RNeasy Plant Mini Kit (Qiagen, Hilden, Germany). Four-week-old sterile seedlings of wild-type plants (ecotype Nossen) were irradiated with 100 Gy of gamma-rays at a dose rate of 300 Gy/h. Six and 12 hours after irradiation, samples were frozen in liquid nitrogen and immediately ground for total RNA isolation. After confirming its quality, extracted RNA was prepared for microarray analysis. An Arabidopsis 2 Oligo Microarray kit (Agilent Technology, CA, USA) was used in this study. After confirming its quality, extracted RNA was prepared for microarray analysis as described by the manufacturer (Agilent) using the suggested chemicals and reagents.