GSM6290616: SERCA2 Knock out [KO2_S58]; Mus musculus; RNA-Seq
Mouse islets were isolated by collagenase digestion. The islets were hand-picked, and allowed to recover overnight in regular RPMI supplemented with 10% FBS, 100U/ml penicillin, and 100μg/ml. RNA was harvested using Qiagen RNAeasy micro kit was used to obtain RNA and the RNA integrity was calciulated using Bio analyzer.  KAPA mRNA Hyperprep kit (KK8581); cDNA library preparation includes mRNA purification/enrichment, RNA fragmentation, cDNA synthesis, ligation of index adaptors, and amplification, following the KAPA mRNA Hyper Prep Kit Technical Data Sheet, KK8581. Each resulting indexed library was quantified and its quality assessed by Qubit and Agilent Bioanalyzer, and multiple libraries pooled in equal molarity. The pooled libraries were then denatured, and neutralized, before loading to Illumina HiSeq 4000 sequencer at 300pM final concentration for 100b paired-end sequencing (Illumina, Inc.). Approximately 30M reads per library was generated.