GSM4065928: WT6m2A; Mus musculus; RNA-Seq
RNA was isolated from flow cytometry-sorted cell populations using RNeasy Micro Kit (74004, Qiagen) A total amount of 10 ng RNA per sample was used as input material for the RNA sample preparations. Libraries were generated using SMART-Seq v4 Ultra Low Input RNA Kit (634892, Takara Bio USA, Mountain View, CA, USA) following manufacturer's recommendations and index codes were added to attribute sequences to each sample. Briefly, first-strand cDNA synthesis from total RNA is primed by the 3' SMART-Seq CDS Primer II A and uses the SMART-Seq v4 Oligonucleotide for template switching at the 5' end of the transcript. PCR Primer II A amplifies cDNA from the SMART sequences introduced by 3' SMART-Seq CDS Primer II A and the SMART-Seq v4 Oligonucleotide for 8 cycles. LD-PCR amplified cDNA is purified by immobilization on AMPure XP beads and then quantified with Agilent Bioanalyzer 2100 system. Approximately 200 pg was used for Nextera XT DNA Library Preparation Kit (Illumina, Cat. Nos. FC-131-1024 and FC-131-1096, San Diego, CA) to make cDNA libraries suitable for Illumina sequencing. Tagmented fragments were amplified for 12 cycles and dual indexes were added to each well to uniquely label each library. Concentrations were assessed with KAPA Library Quantification Kits (KK4844, KAPA, Biosystems, USA) and samples were diluted to ~2 nM and pooled. Pooled libraries were sequenced on the Illumina Novaseq platform and 150 bp paired-end reads were generated.
GEO Accession
GSM4065928