GSM6005304: CD34 Rep2 Multiome (RNA); Homo sapiens; RNA-Seq
GSM6005304
Vial was removed from the storage and immediately thawed at 37 °C in a water bath for 2 min while gently shaking. Next, vial content (1 mL) was transferred to a 50-mL conical tube. To prevent osmotic lysis and ensure gradual loss of cryoprotectant, 1 mL of warm medium (IMDM with 10% FBS supplement) was added dropwise after washing the storage vial, while gently shaking the tube. ingle Cell Multiome ATAC + Gene Expression was performed with a 10X genomics system using Chromium Next GEM Single Cell Multiome Reagent Kit A (catalog no. 1000282) and ATAC Kit A (catalog no. 1000280) following Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Kits User Guide and demonstrated protocol - Nuclei Isolation for Single Cell Multiome ATAC + Gene Expression Sequencing. Briefly, 200k-300k cells (viability 95%) were lysed for 4min and resuspended in Diluted Nuclei Buffer (10x Genomics, PN- 2000207). Lysis efficiency and nuclei concentration was evaluated on Countess II automatic cell counter by trypan blue staining.  9,660 and 16,100 nuclei were loaded per transposition reaction for CD34+ and T-cell depleted marrow, targeting recovery of 6,000 and 10,000 nuclei after encapsulation. After transposition reaction nuclei were encapsulated and barcoded. Next-generation sequencing libraries were constructed following the User Guide, which were sequenced on an Illumina NovaSeq 6000 system.