GSM8946286: Islet, bulk RNA-seq, insulin-depleted, short-read, bio replicate 2; Mus musculus; RNA-Seq
Pancreatic islets were isolated from mice under ketamine/xylazine/acepromazine anesthesia by collagenase delivery into the pancreas via injection into the bile duct. The collagenase-inflated pancreas was surgically removed and digested. After isolation, islets were dissociated using Accutase in a 37°C bead bath for 25-30 minutes. Single-cell suspension was filtered through a 40 mm filter and quenched in RPMI media with 10% FBS. Single-cell suspensions were loaded into a Genomics Chromium targeting 4000 cells per sample. RNA was purified using the Qiagen RNeasy micro kit (Qiagen Cat # 74004) Bulk RNA-seq libraries were prepared using KAPA mRNA Hyper Prep kit (Roche 8098123702), following the KAPA mRNA HyperPrep Kit protocol (KR1352 – v7.21). Insulin depletion was applied between step 8 and 9 in the KAPA library prep protocol.  PacBio MAS-IsoSeq protocol from Jumpcode Genomics. Using the DepleteXTM RNA Depletion Panel (Insulin) kit from Jumpcode Genomics.