RNA-Seq_lscc_pdo
Total RNA was extracted from vehicle-treated or ReACp53-treated (5 M, 144 hours) F-PDOs by using PureLink RNA Mini Kit (Thermo Fisher Scientific).  mRNAs were collected by using NEBNext Poly(A) mRNA Magnetic Isolation Module (New England Biolabs, Ipswich, MA, USA), and RNAseq libraries were prepared by using NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs).  Samples were sequenced on a NovaSeq 600  (Illumina, San Diego, CA, USA) with paired-end 150 base sequencing reads  at 4 Gb per sample.  Sequenced data were tested for quality with FastQC v0.11.8 (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/) and MultiQC v1.12 (http://multiqc.info/).  Reads were quality filtered by using Trimmomatic v0.39 ([44]; http://www.usadellab.org/cms/?page=trimmomatic) and PRINSEQ-lite  v0.20.4 ([45]; https://multiqc.info/).  Reads were then aligned to the human reference genome hg38 by using Spliced Transcripts Alignment  to a Reference (STAR) v2.7.9a ([46]; https://github.com/alexdobin/STAR) and were summarized by using featureCounts v1.6.0, which is part of the Subread package ([47]; https://subread.sourceforge.net/).