GSM4200481
GSM4200481: MM199-Buccal-WES; Homo sapiens; OTHER
Research bone marrow samples were obtained from MM patients at The Princess Margaret Cancer Centre at the time of routine clinical biopsy following informed consent in compliance with the Declaration of Helsinki and a protocol approved by the University Health Network Research Ethics board. Cells were collected in heparinized tubes and processed by Ficoll-Plaque gradient. Plasma cells and B cells were enriched on a Beckman Coulter FACSAria by FSC-SSC characteristics, negative selection for duplets (by FSC and SSC height versus area), negative selection for lineage markers (CD14, CD2, GYPA) and non viable cells (7AAD); and by positive selection for CD38, CD138 or for CD20, as described1. Cell viability was >95% post FACS. Single cell RNA-sequencing libraries were prepared from FACS-purified plasma cells and B cells (pooled) using the 10X Genomics Chromium Single Cell 3' or 5' library performed by Illumina Hiseq 2500 & Gel bead kit v2 and Chromium Single Cell Controller according to the user guide, targeting 3-6,000 cells per sample. Library constituents incorporated a 26bp read 1, 98bp read 2 and an 8bp sample index. Whole exome sequencing libraries were prepared from FACS-purified bone marrow plasma cells as well as from buccal swab cells. DNA was sheared using a Covaris instrument, and 200ng of the sheared product was used for library preparation utilizing the SureSelectXT low input reagent kit. Library quantity and quality were verified by Bioanalyzer. Whole exome sequencing was performed using an Illumina Hiseq 2500.
GEO
GSM4200481
GEO Accession
GSM4200481