GSM8033107: K9TCC_Mx, H3K27ac; Canis lupus familiaris; ChIP-Seq
Cells were harvested using a Cell Lifter in PBS, pelleted and lysed in three stages with a protease inhibitor cocktail (Roche #11873580001) at each step. DNA in pelleted nuclei was sheared on ice using a GEX130 Ultrasonic processor in 3 mL volumes at 30% output, 25s ON, then 60s OFF; repeated 8X. MagBead previously bound with antibody with histone modification washed with PBS, resuspended and added to fragmented DNA and incubated at 4C o/n. Beads with bound DNA strands were washed 5X, rinsed with PBS. Final rinse was aspirated and 115 uL of elution buffer was added and incubated at 65C for 15 minutes vortexing every 2 min followed by an o/n incubation on a Thermomixer R. Libraries were constructed using paired-end libraries with inserts averaging 320 bps and run on an Illumina HiSeq 2000 with 100 bp reads.