Integrated genomic and transcriptomic analysis improves disease classification and risk stratification of MDS with ring sideroblasts

CD34+ cells were isolated from the MNC using AUTO-MACS with double-separation option (Miltenyi Biotec, Germany) and submitted for RNA extraction. RNA was extracted with RNeasy Microkit (Qiagen, Hilden, Germany) and treated with DNase, according to manufacturer instruction.

RNA integrity number was estimated using Agilent RNA 6000 Pico (Agilent Technologies, Palo Alto, CA) and was greater than 6.5 for all the samples (median 8.2). The RNA-sequencing (RNA-seq) libraries were prepared from total RNA using SMARTer Stranded Total RNA-Seq Kit v2 Pico Input Mammalian with enzymatic ribosomal depletion (Takara Bio, Japan). Libraries were sequenced using the Novaseq 6000 with paired-end 150bp configuration.