Gene mutation analysis was performed using DNA extracted from bone marrow aspirate specimens in a small subset of patients. Amplicon-based next generation sequencing (NGS) targeting the entire coding regions of a panel of 81 genes was performed using a MiSeq platform (Illumina, San Diego, CA, USA) to detect somatic mutations and insertions and/or deletions as previously described [11 (link)]. The 81-gene panel included: ANKRD26, ASXL1, ASXL2, BCOR, BCORL1, BRAF, BRINP3, CALR, CBLB, CBLC, CBL, CRLF2, CREBBP, CEBPA, CSF3R, CUX1, DDX41, DNMT3A, EED, ELANE, ETNK1, ETV6, EZH2, FBXW7, FLT3, GATA1, GATA2, GFI1, GNAS, HNRNPK, HRAS, IDH1, IDH2, IKZF1, IL2RG, IL7R, KRAS, JAK2, JAK3, KDM6A, KIT, KMT2A, MAP2K1, MPL, NF1, NOTCH1, NPM1, NRAS, PAX5, PHF6, PIGA, PML, PRPF40B, PTEN, PTPN11, RAD21, RARA, RUNX1, SETBP1, SF1, SF3A1, SF3B1, SH2B3, SMC1A, SMC3, SRSF2, STAG1, STAG2, STAT3, STAT5A, STAT5B, SUZ12, TERC, TERT, TET2, TP53, U2AF1, U2AF2, WT1, and ZRSR2. All coding exons for each gene were covered with an analytical sensitivity of 5% mutant reads in a background of wild-type reads. Established bioinformatics pipelines were used to identify somatic variants.
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