GSM8741363: Rat embryonic cortical cells at E18; Rattus norvegicus; RNA-Seq
RNA extraction was integrated in "library construction protocol" process. Whole-cell suspensions harvested from the expansion culture were pelleted and were subjected to Cell Multiplexing Oligo labeling.  Alternatively, samples for analysis were cryopreserved as follows: whole-cell suspensions harvested from the expansion culture were pelleted and were resuspended in Cell Banker Type 1 Plus for cryopreservation at −80˚C.  For Cell Multiplexing Oligo labeling, after thawing in the water bath at 37℃ if needed, collected cells were resuspended in Cell Multiplexing Oligo solution and were incubated for 5 minutes at room temperature.  After the incubation, cells were washed with 1% BSA (SIGMA, A1519)-PBS (−) and were collected by centrifugation at 200g for 10 minutes.  Collected cells were resuspended in 1% BSA-PBS (−) containing anti-TRA-1-85 antibody (BD Bioscience, 563302, 1:20) and placed on ice for 30 minutes in the dark.  Then, the cells were washed with PBS once and were resuspended in 1% BSA-PBS (−) containing 3 μM DRAQ7 (abcam, ab109202).  To isolate live human cells, TRA-1-85 (+) DRAQ7(−) cells were sorted into DMEM/F12 (GIBCO, 10565-018) + 1% BSA by FACS Aria III.  Sorted cells were subjected to the subsequent library construction using 10X Genomics Single Cell 3' Kit v. 3.1, followed by sequencing on an Illumina NovaSeq 6000 platform.  All steps were performed in accordance with the manufacturer's instructions.