GSM1545901: Dsim_Dam_rep2; Drosophila simulans; ChIP-Seq
To extract high-molecular weight genomic DNA, each aliquot of embryos was dechorionated and  homogenized  in a Dounce 15-ml homogenizer in 10 ml of homogenization buffer (10 mM Tris-HCl pH 7.6, 60 mM NaCl, 10 mM EDTA, 0.15 mM spermine, 0.15 mM spermidine, 0.5% Triton X-100). The lysate was then  spun for 10 minutes at 6000g. The supernatant was discarded, and the pellet was  resuspended in 10 ml homogenization buffer, then spun again for 10 minutes at  6000g. The supernatant was again discarded, and the pellet was resuspended in 3  ml homogenization buffer. 300 μl of 20% n-lauroyl sarcosine were added, and the  samples were inverted several times to lyse the nuclei. The samples were treated  with RNaseA followed by proteinase K, then purified by two  phenol-chloroform extractions and one chloroform extraction. DamID samples were prepared from genomic DNA following the DamID protocol from Vogel et al. (Nat Protoc 2, 1467-1478, 2007). Libraries were prepared as for ChIP-seq by BGI Tech, starting with at least 20 ng of DNA per sample and following standard Illumina protocol (end repair, 3' dA-tailing, ligation of adapters, PCR amplification and size selection for fragments 100-300bp). Libraries were multiplexed with 2 samples per run for the MiSeq and 9-12 samples per lane for the HiSeq. MiSeq libraries were run as 150-bp single-end reads, while HiSeq libraries were run as 50-bp single-end reads.