GSM2092590: Crz1_A_T1; Saccharomyces cerevisiae; ChIP-Seq
Forty ml of cultures were taken at the appropriate times and made 1% formaldehyde to crosslink proteins and DNA. From this point the procedure was as described in (González et al., 2013;Serra-Cardona et al., 2014) with the following modifications: i) chromatin was fragmented using a Bioruptor Plus UCD-300 equipment (Diagenode) provided with a cooling system (4°C) for 30 cycles (high intensity; 30 s of sonication followed by 60 s pause) to generate fragments of ≤ 500 bp length; ii) the supernatant was pre-cleared with protein G sepharoseTM fast flow (GE Healthcare, #17-0618-01) beads for 1 h at 4°C and then, the pre-cleared cell lysate was incubated with 1 µg of polyclonal anti-HA ChIP-grade (Abcam, #ab9110) antibody overnight at 4°C; iii) anti-HA-Crz1-DNA complexes were collected with protein-G-sepharose beads incubating for 1 hour at 4°C,  and iv) after reverse crosslinking, DNA was extracted using a NucleoSpin® Gel and PCR Clean-up kit (Macherey Nagel, #740609) and NTB buffer (Macherey-Nagel, #740595). ChiP libraries were prepared using the TruSeq ChIP Sample Preparation Kit (Illumina) and then subjected to paired-end deep sequencing.
GEO Accession
GSM2092590