title: GSM9268837: CD271+ MSC Plate3; Homo sapiens; RNA-Seq
library_construction_protocol: CD45-CD14-CD235a-CD71-CD38-CD271+ single cells were sorted in 5 384-well plates containing 5 μl of CEL-Seq2 primer solution in mineral oil (24 bp polyT stretch, a 4 bp random molecular barcode (UMI), a cell-specific barcode, the 5'-Illumina TruSeq small RNA kit adapter, and a T7 promoter), provided by Single Cell Discoveries (Utrecht, the Netherlands). After sorting, the plates were centrifuged for 1 min at 1000g and immediately placed on ice and stored at −70°C. Single Cell Discoveries further processed the plates. In brief, ERCC Spike-in RNA Mix (Ambion, 0.02 μl of 1:50,000 dilution) was added to each well before cell lysis with heat shocking. Reverse transcription and second-strand synthesis reagents were dispensed using the Nanodrop II (GC biotech). After generation of cDNA from the original mRNA, all cells from one plate were pooled and the pooled sample was amplified linearly with in vitro transcription. To generate sequencing libraries, RPI-series index primers were used for library PCR. Libraries were sequenced on an Illumina Nextseq500 using 75 bp paired-end sequencing.