GSM2571732: P10-1_ATAC-seq; Mus musculus; ATAC-seq
Both retinas from mice age P3, P6, and P9 were used for the ATACseq experiment. For each time point, three mice were used. Only male mice were used to avoid any influence from X-chromosome inactivation. A protocol from Siegert et al. (Nature Neuroscience 2012) was used to dissociate retinal cells. Retinas were dissected in HBSS and incubated in an activated papain solution for 5 min at 37 °C. After removing the papain solution, retinas were washed with HBSS containing 2% FCS and then dissociated in HBSS containing 2% by triturating the tissue with a fire-polished Pasteur pipette. The suspension was filtered before FACS sorting. Single-cell suspensions were obtained after FACS sorting into PBS/10%FCS. ATACseq was performed according to Buenrostro et al. (Nature Methods, 2013), with minor modifications. Briefly, 50,000 cells were used per transposition reaction. Cells were collected and lysed in cold lysis buffer (10mM Tris-HCl pH 8.0; 10mM NaCl; 3mM MgCl2; 0.5% NP40).  Transposition was done as reported in Buenrostro et al. (Nature Methods, 2013), steps 6 – 12, and PCR amplification was done according to steps 13 – 18. Briefly, during the amplification reaction, PCR primers were used as mentioned in the supplemental material from Buenrostro et al. (Nature Methods, 2013). Amplified libraries were cleaned up using AMPure XP beads in a DNA:beads ratio of 1:1.8. Further size selection of the library prior to sequencing was avoided. The Nextera DNA Library Preparation Kit was used for library preparation. Multiple experiments were sequenced per lane (up to 3 per lane). Libraries were sequenced paired-end 100 bp reads using the Illumina HiSeq 2500.