Illumina HiSeq 2500 paired end sequencing; Distinct and sexually dimorphic X dosage compensation states in the mammalian germline
Distinct and sexually dimorphic X dosage compensation states in the mammalian germline
Timed matings were undertaken, and noon on the date of the vaginal plug was taken to be E0.5. Embryos were subsequently harvested  on the stipulated day. Germ cells (GFP-positive) and associated somatic cells (GFP-negative) were isolated separately from individual embryos using FACs sorting on the MoFLo XDP or FACS Aria platforms. Live cells, i.e. only those staining negative for propidium iodide, were collected, and typically purity checking on the GFP negative populations was >99%. RNA was isolated from FACs sorted purified cell populations using the Ambion PicoPure RNA isolation kit. Eluted RNA was used to obtain double-stranded-cDNA using the Clontech (SMARTER) Ultralow input RNA-Seq Kit according to the  manufacturer’s protocol, or the Smart-Seq2 protocol. cDNA was normalized to 10ng in low-TE buffer and was sheared using Covaris S-series, and was cleaned up with Zymo DNA conc-5 and eluted in 12ul low TE. Libraries of ~300bp were generated   510 using Nugen Ovation® Ultralow DR (part No. 0330) with 15 PCR cycles.
Turner Group
E-MTAB-4616:Sample 76
Experimental Factor: XY Male
genotype
Experimental Factor: Somatic cells (gonadal)
cell type
Experimental Factor: embryonic day 14.5
developmental stage
Experimental Factor: 1
replicate