GSM2200164: Zta_CHIPSeq_2; Homo sapiens; ChIP-Seq
Chromatin was fixed by application of 1% (v/v) formaldehyde to cells for 10min, at 20oC and was extracted as described in Bark-Jones et al. (2006). A Vibra-Cell (SONICS) was used to shear the chromatin. HONE1-EBV cell lysates were subjected to sonication in a volume of 100μl on ice for ten rounds, with 10s pulses and a 21% amplitude output. Samples were mixed after every minute. The efficacy of sonication was assessed by analysing the sample on a 1% (w/v) agarose gel. Chromatin from 3.5x10^7 cells was used to immunoprecipitate Zta-associated-DNA using 10ug EBV ZEBRA antibody (sc-53904) (Santa Cruz). Immune complexes were collected with 50% pre-blocked protein A/G-sepharose beads. Following wash steps and protein digestion of immune complexes, eluted DNA was purified using a PCR purification kit (Qiagen) and eluted in 45μl. The DNA libraries were prepared according to the NEBNext ChIP-Seq Library Prep Reagent Set for Illumina kit (New England Bioscience), switching the PCR and the size selection steps. Index 4 was used for the Input, Index 6 for ChIP1 and Index 12 for ChIP2. Size selection was performed on a 2% (w/v) agarose gel with SYBR Gold dye to exclude the adaptor from the sequencing process and to purify DNA fragments in the 175-225bp size range. Quality control was performed using the Agilent 2100 Bioanalyzer.
GEO
GSM2200164