title: GSM3466902: 10X 2675 retinal single-cells batch F2; Mus musculus; RNA-Seq
library_construction_protocol: Coordinated pregnant mice of 12 to 30 weeks were ethically sacrificed to extract E15.5 embryos eyes. Thirty retinas were extracted in ice-cold L15, micro-dissected under a stereomicroscope and incubated in 200µl single cell dissociation solution consisting in a papain (1mg/ml) enriched HBSS at 37°C for 12 minutes with trituration every 2 minutes. At the end of the incubation time, cells were further dissociated via gentle up-and-down pipetting. Reaction was stopped with the addition of 400µl of ovalbumin enriched cold HBSS and the cell suspension was then passed on a 40µm cell strainer to remove cellular aggregates.  Cells were then centrifuged 5 min at 500 G at 4°C. After removal of the liquid, the pellet was suspended in 250µl of cold HBSS and the resulting solution was finally FAC-sorted on a MoFloAstrios device (Beckman) to reach a concentration of 410 cells /µl. The libraries of single-cells were prepared using the Chromium 3' v2 (chimistry SC3pv2) platform following the manufacturer's protocol (10X Genomics, Pleasanton, CA). Briefly, single cells were partitioned into Gel beads in EMulsion (GEMs) in the GemCode instrument followed by cell lysis and barcoded reverse transcription of RNA, amplification, shearing and 5' adaptor and sample index attachment. On average, approximately 7,000 single cells were loaded on each channel with 2675 cells recovered for the first replicate (index F2), and 2673 cells recovered in the second (index E2). Libraries were sequenced on the Illumina HiSeq 4000 (Paired end reads: Read 1, 26bp, Read 2, 98bp).
attributes.tag: GEO Accession
attributes.value: GSM3466902