概要: Here we present single-cell RNA sequencing (scRNA-seq) data of two LCLs: GM12878 and GM18502, produced from the blood of two female donors with European and African ancestry, respectively.
项目整体设计: Cells of three samples (the two LCLs and a 1:1 mixture of the two) were prepared separately using a 10X Genomics Chromium Controller and deeply sequenced.
GM12878 and GM18502 were purchased from the Coriell Institute for Medical Research. Cells were cultured in the Roswell Park Memorial Institute (RPMI) Medium 1640 supplemented with 2mM L-glutamine and 20% of non-inactivated fetal bovine serum in T25 tissue culture flasks. Flasks with 20 mL medium were incubated on upright position at 37℃ under 5% carbon dioxide. Cell cultures were split every three days for maintenance.
处理方案:
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提取方案:
Single-cell sample preparation was conducted according to Sample Preparation Demonstrated Protocol provided by 10X Genomics as follows: 1 mL of cell suspensions from each cell line (day 4, stable phase) was pelleted in Eppendorf tubes by centrifugation (400g, 5min). The supernatant was discarded, and the cells pellet was then resuspended in 1X PBS with 0.04% BSA, followed by two washing procedures by centrifugation (150g, 3min). After the second wash, cells were resuspended in ~500 uL 1X PBS with 0.04% BSA followed by gently pipetting mix 10-15 times.
建库方案:
Libraries were prepared using the Chromium Controller (10X Genomics, CA) in conjunction with the single-cell 3'v2 kit. Briefly, the cell suspensions were diluted in nuclease-free water according to manufacturer instructions to achieve a targeted cell count of 5,000 for each cell line. The cDNA synthesis, barcoding and library preparation were then carried out according to the manufacturer's instructions. Libraries were sequenced in the North Texas Genome Center facilities on a Novaseq 6000 sequencer (Illumina, San Diego).
