title: GSM5252334: AMLMRD004-Dx; Homo sapiens; RNA-Seq
library_construction_protocol: Single-cell RNA sequencing was performed following the Drop-seq Online Protocol by Macosko et al (http://mccarrolllab.com/dropseq/) with minor modifications. Briefly, single cell suspension were loaded into the microfluidic device (FlowJEM) together with droplet-generating oil (Bio-Rad) and barcoded beads (MACOSKO-2011-10, Chemgenes) suspended in lysis buffer supplemented with RNase inhibitor (Lucigen) at a final concentration of 1 U/µL. Emulsions were incubated on ice for 30 minutes before droplet breakage and bead collection. Beads were subject to reverse transcription (Maxima H Minus Reverse Transcriptase: ThermoFisher; Template switch oligo: IDT) and Exo I (New England Biolabs) digestion. Next, 4000 beads were aliquoted into one PCR tube for amplification (KAPA HotStart ReadyMix: Kapa Biosystems). Products from multiple parallel PCRs were pooled together and purified using AMPure XP Beads (Beckman Coulter Life Sciences). For each sample, 1 ng high quality cDNA was used for tagmentation and NGS library preparation using a Nextera XT kit (Illumina). High quality NGS libraries were sequenced on an Illumina NextSeq 550 (75-cycle kit) or NovaSeq 6000 (100-cycle kit) sequencers. All paired AML samples were processed and sequenced together to minimize batch effects.
attributes.tag: GEO Accession
attributes.value: GSM5252334