GSM4386205: WT_single-cell RNA-seq; Mus musculus; RNA-Seq
After collection of cells , centrifuge at a speed not exceeding 400rcf (adjust the speed and centrifugation time according to the cell types ). Carefully discard the supernatant and retain the cell precipitation.  The cell pellet was resuspended by 1mL 1X PBS containin g 0.04% BSA and the washing procedure was repeate d twice. After washing, appropriate volume PBS was added to the cell precipitation to obtain single cell dispersion suspension with a concentration c lose to  the goal number. A type of wide bore pipette tip was used in cell resuspension for a lower cell dam age. If necessary, cell filter should be applied to remove cell debris and clusters ( eg. 30 µm MACS SmartStrainers and 40 µm Flowmi TM Cell Strainer).  Automatic cytometry was used to determine the cell concentration. And the sample volume was calcul ated based on the optimal cell sampling concentration supplied by 10X official website and the target capture number . If the calculated concentration is too high, adjust the liquid volume to the appr opriate concentration and repeat counting once more. Once the desired cell suspension was obtained , immediately place the samples on ice for subsequent GEMs preparation and reverse transcription. RNA libraries sequencing  was performed using standard Illumina protocols. 1) Interrupt, end repair and add A base. Finally the SPRIselect beads was used to purify the product. 2) Adaptor ligation and SPRIselect purification. 3) Index PCR and SPRIselect purification. 4) Qubit® 3.0 Flurometer Life Technologies, CA, USA  was used to determine the library  concentration.
GEO Accession
GSM4386205