title: GSM8790230: PBMC_DMSO_1; Homo sapiens; RNA-Seq
library_construction_protocol: Bulk-seq: RNA was extracted by trizol method (Invitrogen, USA), RNA integrity was assessed using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system (Agilent Technologies, CA, USA). 1.3 ug of total RNA was used for the construction of sequencing libraries. sc-RNA-seq: The cell suspension was loaded into Chromium microfluidic chips with 3' v2 chemistry and barcoded with a 10× Chromium Controller. RNA from the barcoded cells was subsequently reverse-transcribed and constructed sequencing libraries. Bulk-seq:Total RNA was used as input material for the RNA sample preparations. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in First Strand Synthesis Reaction Buffer(5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase(RNase H-). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3' ends of DNA fragments, Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 370~420 bp in length, the library fragments were purified with AMPure XP system. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system. sc-RNA-seq:Sequencing libraries constructed with reagents from a Chromium Single Cell 3' v2 reagent kit according to the manufacturer's instructions.