Pediatric sAML (from JMML) #1
Cryopreserved bone marrow samples were thawed and the number of cells and their viability were determined using a Countess Automated Cell Counter (Thermo Fisher Scientific, USA). A cell suspension with viability greater than 80% was diluted in cell buffer to a concentration of 3000 to 4000 cells/l in a total volume of 50 l for loading into a Tapestri microfluidic cartridge. Single-cell DNA libraries were generated with Mission Bio Tapestri Single-ell DNA Myeloid Kit (Mission Bio, USA) according to the manufacturers instructions. Targeted exons and splice sites for the genes are presented in the appendix 2. DNA libraries were sequenced using a NextSeq 550 instrument (Illumina, USA) with 150-bp paired-end reads and 60-80X sequencing depth. The fastq files were processed with Tapestri Pipeline (Mission Bio, USA) for read alignment to the reference genome, filtering, barcode counting, and normalization.