title: RNA-seq of mouse primary intestinal fibroblasts (Lkb1fl/fl AdCre or AdGFP) treated with or without LPS
design: RNA-seq of mouse primary intestinal fibroblasts (Lkb1fl/fl AdCre or AdGFP) treated with or without LPS
library_construction_protocol: primary intestinal fibroblasts derived from Lkb1fl/fl mice were treated with either AdCre or AdGFP adenovirus and at day 4 after transduction treated with or without LPS (1ug/ml). Cells were lysed and RNA extracted after 24h. RNA from fibroblasts was isolated using the Nucleospin RNA II isolation kit (Macherey-Nagel)  RNA sequencing was performed using 3â€²UTR RNA sequencing with a 'Pool Seq' method, which is based on the Dropseq approach for single-cell sequencing (Macosko et al., 2015). The method involves priming mRNA using an oligo-dT primer incorporating a 12 bp barcode and an 8 bp unique molecular identifier (UMI) sequence. The UMI enables the removal of PCR duplicates during data analysis. To generate double-stranded cDNA, the single-stranded cDNA was converted using the template-switch effect, followed by PCR amplification using SMART PCR primers.