Illumina MiSeq sequencing; ChIP-exo of S.cerevisiae transcription factors Fkh1 and Fkh2 under logarithmic growth and stationary
ChIP-exo of S.cerevisiae transcription factors Fkh1 and Fkh2 under logarithmic growth and stationary
1. Grow on YPD+G418 plates 2. Pick colony mid-day into 75ml 1/2 diluted delft+CSM+2%glu 3. Next morning, start mid-log phase culture, 4x 250ml shakeflasks with 150ml 1/2 diluted delft+CSM+2%glu. Add to make starting OD 0.2. Leave ~5h before checking OD. Target 0.8. ON cultures are left shaking until afternoon. ChIP-seq on beads: Pierce Anti-c-Myc Magnetic Beads (88843).  1. From OD, calculate how to get 3x33 OD into 3x50ml tubes. Prepare water with 37% formaldehyde 1:36, 1.4ml to each 50ml tube then add appropriate amount of cultures to get 33OD. Shake on tipping board RT 15min 2. Add 2.5M glycine to a final concentration of 125mM (1:19), 2.6ml to 50ml tubes, leave on shaker 5min 3. Pellet cells 1500g 4c 3min. Wash once in 20ml then once in 10ml ice-cold TBS 4. After final wash, move to eppi with small remaining amount of liquid. Get box with liquid N2 and drop tubes in. Move to -80c for storage   ChIP-exo library preparation as described in manuscript