GSM9401972: LPMIC011_RNA; Homo sapiens; RNA-Seq
For RNA-seq library prepration, PDX tumors were homogenized, and total RNA was extracted using the AllPrep DNA/RNA Kit (Qiagen, 80204) and quantified with the Qubit High Sensitivity RNA Kit (Life Technologies, Q32852). One microgram of RNA was used to generate sequencing libraries. For ATAC-seq library prepration, viably cryopreserved PDX tumor cells were thawed and processed using the Dead Cell Removal Kit to eliminate non-viable cells. Fifty thousand live tumor cells were washed in ATAC-seq resuspension buffer, resuspended in 50 μL of ATAC-seq resuspension buffer containing 0.1% NP-40, 0.1% Tween-20, and 0.01% digitonin, and incubated on ice for 3 min. Following lysis, nuclei were washed with cold ATAC-seq resuspension buffer containing 0.1% Tween-20 (without NP-40 or digitonin), then resuspended in 50 μL of transposition mixture and incubated at 37 °C for 30 min in a thermomixer at 1000 rpm. Fragmented DNA was purified using the Zymo DNA Clean & Concentrator-5 kit (Zymo, D4024). For RNA-seq library prepration, one microgram of RNA was used to generate sequencing libraries with the KAPA HyperPrep RNA Kit with RiboErase (Roche, KK8560), using 8 cycles of PCR amplification according to the manufacturer's instructions. For ATAC-seq library prepration, libraries were prepared with the KAPA HyperPrep Kit (Roche, KK8502) using 10 cycles of PCR amplification. Both RNA-seq and ATAC-seq libraries were assessed on a TapeStation 4200 (Agilent, G2991BA), quantified with the Qubit High Sensitivity dsDNA Kit (Life Technologies, Q32854), multiplexed, and sequenced on an Illumina NovaSeq 6000 using a 100-10-10-100 bp read configuration.