GSM5989998: Young rep 3; Mus musculus; RNA-Seq
For RNA sequencing, RPMs (live, single, CD45+, B220-, GR1-, F4/80-high, CD11b-dim, TREML4+) were sorted into Trizol-LS (Invitrogen, 10296010). RNA was isolated using Direct-zol RNA Microprep Kit (Zymo Research, R2062) according to the manufacturer's instructions. To prepare libraries from FACS-sorted RPMs (at least 100 000 cells/sample) we used the previously described Smart-seq2 protocol {Picelli, 2013 #1236}, suitable for low-input total mRNA sequencing. The quality of RNA and material during the preparation of libraries was checked by Bioanalyzer. . The samples were sequenced on NextSeq500 (Illumina) with 75 bp single-end reads, with ~50 million reads/sample. RNAseq was performed at GeneCore at EMBL (Heidelberg, Germany)