Trueseq small rna library preparation

RNA was isolated using the RNeasy Plus Kit (Qiagen), and the purity of total RNA per sample was verified using the Agilent Bioanalyzer (Agilent Technologies, Santa Clara, CA). RNA sequencing was performed through the Yale Center for Genome Analysis using an Illumina HiSeq 2000 platform (paired-end 150 bp read length). Briefly, rRNA was depleted from RNA using Ribo-Zero rRNA Removal Kit (Illumina). RNA libraries were generated from control cells using TrueSeq Small RNA Library preparation (Illumina) and sequenced for 45 cycles on Illumina HiSeq 2000 platform (paired-end, 150 bp read length).

RNA was isolated using the RNeasy Plus Kit (Qiagen) and purity of total RNA per sample was verified using the Agilent Bioanalyzer (Agilent Technologies, Santa Clara, CA). RNA sequencing was performed through the Yale Center for Genome Analysis using an Illumina HiSeq 2000 platform (paired-end 150 bp read length). Briefly, rRNA was depleted from RNA using Ribo-Zero rRNA Removal Kit (Illumina). RNA libraries were generated from control cells using TrueSeq Small RNA Library preparation (Illumina) and sequenced for 45 cycles on Illumina HiSeq 2000 platform (paired end, 150 bp read length).

Total RNA from unstimulated BMDMs was extracted as described above, and RNA was purified using and miRNA isolation Kit (Qiagen) followed by DNase treatment to remove genomic contamination using RNA MinElute Cleanup (Qiagen). The purity and integrity of total RNA samples were verified using the Agilent Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). rRNA was depleted from RNA samples using Ribo‐Zero rRNA Removal Kit (Illumina). Small RNA libraries from WT BMDMs were performed using TrueSeq Small RNA Library preparation (Illumina) and were sequenced for 45 cycles on Illumina HiSeq 2000 platform (2 × 50 bp read length). MicroRNA sequencing results were trimmed and mapped to miRBase mouse stem‐loop sequences (http://www.mirbase.org/) using the Bowtie alignment (http://bowtie-bio.sourceforge.net/index.shtml) program (version 0.12.7). The alignment reads were normalized as proportion of total reads mapped to any know miRNAs in each sample. The data discussed in this publication have been deposited in NCBI Gene Expression Omnibus and are accessible through GEO Series accession number GSE97622 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE97622).