GSM3993358: GFP+ ILCPs from Rag1cre;ET-2 rep2; Mus musculus; RNA-Seq
ILCps were directly sorted into TRIZOL. Trizol samples were processed using Zymo Research kit RNA Clean & Concentrator (Cat: R1015S) Zymo Research kit Direct-zol RNA MicroPrep (Cat: R2060) An mRNA library was generated from extracted total RNA using the Trio RNA-seq kit from NuGEN. Briefly, extracted total RNA underwent DNase treatment followed by double stranded cDNA generation. cDNA was first purified using Agencourt RNAClean XP beads and then further amplified using Single Primer Isothermal Amplification (SPIA). Following end repair, adapter ligation, and amplification of properly ligated library molecules the library underwent depletion of rRNA molecules using the NuGEN AnyDeplete system, a probe-based enzymatic digestion of unwanted molecules. Remaining library molecules were amplified again and quantified on an Invitrogen Quantseq fluorometer followed by sizing on an Agilent Tapestation. After normalizing dilutions the samples were pooled as equal volumes and sequenced on an Illumina NextSeq 500 with High Output chemistry giving paired end 75bp reads.