GSM8229690: GSA_biol_rep_2.1; Homo sapiens; RNA-Seq
200 hematopoietic progenitor cells (CD34+, CD43+, CD33+ and DAPI -) derived from parental and mutant hiPSC were sorted at day 15 of EB differentiation directly in 8µl lysis buffer to 0.2% Triton X-100, and 2 U/μL RNase inhibitor (e.g., SUPERase•In, Thermo Fisher Scientific) Reverse Transcription (RT) and cDNA Amplification: The released RNA was reverse transcribed into complementary DNA (cDNA) using the Smart-seq2 protocol. Briefly, a template-switching oligo (TSO) (5'-AAGCAGTGGTATCAACGCAGAGTACATrGrG+G-3') was annealed to the RNA molecules, and reverse transcription was performed using SuperScript II reverse transcriptase (Invitrogen) at 42°C for 90 minutes, followed by 10 cycles of 50°C for 2 minutes and 42°C for 2 minutes. The cDNA was then purified using AMPure XP beads (Beckman Coulter) and amplified using KAPA HiFi HotStart ReadyMix (Kapa Biosystems) with 18 cycles of PCR amplification Prepared cDNA libraries were generated using Nextera Low Input library preparation kit for whole transcriptome sequencing on the HiSeq2000