GSE169348
Next Generation Sequencing Facilitates Quantitative Analysis of IL4/IL13 treated wild-type BMDMs
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived IL4/IL13 treated BMDMs transcriptome profiling (RNA-seq) to quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods. Methods: BMDMs mRNA profiles of 8 weeks old wild-type (WT) mice were generated, stimulated with IL4/IL13, IL4/IL13+ TNF or TNF, deep sequenced, in triplicate, using Illumina NovaSeq 6000 platform. Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9). Conclusions: Our study represents the first detailed analysis of IL4/IL13 or IL4/IL13+TNF stimulated BMDMs, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions. Overall design: mRNA profiles of wild-type BMDMs stimulated with IL4/IL13, IL4/IL13 + TNF or TNF for 6 or 24 hours