RNA-seq analysis of mouse lung tumors

Tumor samples were micro-dissected away from normal lung tissue and RNA was extracted using Qiagen RNeasy Micro kit per manufacturers instructions. Sequencing libraries were prepared on the Illumina NeoPrep and subjected to 75 bp paired end sequencing on the Illumina NextSeq 500 platform. Fastq files for each sample were aligned against the mouse genome, build GRCm38.p4, using the STAR aligner (v2.5.2b) [25 (link)]. FeatureCounts (v1.5.0-p1) was used to quantify alignments against the mouse genomic annotations from Gencode (vM11) [26 (link)]. High purity of tumor cell derived RNA was estimated by detecting SNPs that were engineered during design of the Kras^LSL-G12D allele in the Integrative Genomics Viewer [27 (link), 28 (link)]. Differentially expressed genes were identified with DESeq2 (v1.14.0) [29 (link)].

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Walter D.M., Venancio O.S., Buza E.L., Tobias J.W., Deshpande C., Gudiel A.A., Kim-Kiselak C., Cicchini M., Yates T.J, & Feldser D.M. (2017). Systematic in vivo inactivation of chromatin regulating enzymes identifies Setd2 as a potent tumor suppressor in lung adenocarcinoma. Cancer research, 77(7), 1719-1729.

Publication 2017

RNeasy Micro kit NeoPrep NextSeq 500

Allele Cell Genes Genomic Lung Mouse Normal tissue Snps Tumor

Corresponding Organization : UPMC Hillman Cancer Center

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Protocol cited in 3 other protocols

Variable analysis

independent variables

Kras^LSL-G12D allele

dependent variables

Differentially expressed genes

control variables

Purity of tumor cell derived RNA
Mouse genome build GRCm38.p4
Gencode vM11 genomic annotations

Annotations

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