GSM9313100: B16_F10_CT_1; Mus musculus; RNA-Seq
RNA was extracted from FFPE sections using the Quick RNA-FFPE kit (Zymo Research catalog #R1008) when tissue samples were limited. Reverse transcription was carried out using the High-Capacity RNA-to-cDNA™ Kit (ThermoFisher catalog #4387406) using between 100ng – 1ug of RNA per the manufacturer's instructions. Following RNA isolation, FFPE total RNA integrity was checked using a 2100 Bioanalyzer with a DV200 reading as 47% - 61% for all six samples (Agilent Technologies, Santa Clara, CA). RNA concentrations were measured using the NanoDrop system (Thermo Fisher Scientific, Inc., Waltham, MA).  We used 200ng of FFPE total RNA sample as the input for fragmentation by treating with metal ions present in the 2X Priming Buffer at elevated temperature (Agilent SureSelect XT HS2 RNA Library Preparation and Target Enrichment Protocol #G9989-90000, Agilent Technologies, Santa Clara, CA). Fragmented RNA was used to perform the first and the second-strand cDNA synthesis. After AmpPure XP Beads purification, cDNA samples proceeded to end repair, A-tailing and adapter ligation following the manufacturer instructions. The adaptor-ligated cDNA library was then PCR amplified and hybridized by SureSelect XT Mouse All Exon probes (Agilent Technologies, Catalog #5190-4641). Captured and target-enriched cDNA libraries were amplified and purified.