DOI: 10.5281/zenodo.3583340
title: Supplementary Data - Nanopore cDNA With and Without mtDNA

description: Update (v1.5.6-countAnalysis): added / updated a script to convert transcript mappings to a count table: count_analysis.r Update (v1.5.5-Int): added intermediate (post-demultiplexing) demonstration dataset: SampleDESet_60k_demultiplexed.tar.gz Also updated count tables and DESeq2 results: raw_counts_2019-Nov-29.csv DE_orig_VST_GRCm38_CG_GL261_2019-Nov-29.csv.gz DE_shrunk_VST_GRCm38_CG_GL261_2019-Nov-29.csv.gz Update (v1.5.4-MT): updated demonstration dataset (now 40Mb) to include mitochondrial transcripts, and exclude non-target sequences from chimeric reads: reads_SampleDESet_sampled_60k.fastq.gz Mm_subset.GRCm38.ensembl_v98.fa Update (v1.5.3-Ensembl): added/updated Ensembl reference transcripts, including non-coding RNA: ensembl_mm10_geneFeatureLocations.txt.gz - genome annotation Mus_musculus.GRCm38.ensembl_v98.fa.gz - transcript sequences Update (v1.5.2-Demo): added 60Mb demonstration dataset for teaching / learning purposes (<em>reads_SampleDESet_sampled_60k.fastq.gz</em>). This dataset can be used for testing demultiplexing, or carrying out differential expression analysis on nanopore reads. There are 15k reads sampled from each run (subset on ~300 genes), plus 7.5k chimeric reads from the Dec17 run, plus 7.5k reads sampled from from CG009/BC08. The gene subset includes ~100 genes differentially-expressed between 4T1WT and 4T1ρ<sup>0</sup> cell lines, plus an additional ~200 genes with minimal differential expression observed between any 4T1 cell lines (i.e. a housekeeping gene set): BC01, BC02, BC04, BC06 - 4T1 cell line reads from three sequencing runs (005, 005, 004, Dec17) BC03, BC05, BC07 - 4T1ρ<sup>0</sup> (without mtDNA) cell line reads from three sequencing runs (005, 004, Dec17) BC08 - 4T1ρ<sup>0</sup>SC (from subcutaneous tumour) cell line reads from one sequencing run (009) [note: v1.5.1-Demo has the wrong sequence ID for BC08 reads] Update (v1.5.0-Called): added full 4T1 call set (called using guppy high-accuracy v3.2.2), gene transcript count tables, and differential expression results Update (v1.4.1-Chimeric): added more example called chimeric reads of various [unclassified] categories Update (v1.4.0-Chimeric): added example chimeric reads, one formed [presumably] during sample prep ["conjugation"], another formed in-silico ["double read"] Update (v1.3.0-DESeq2): added count table and metadata file for DESeq2 analysis Update (v1.2.1-10k): added 10k of FASTQ reads from a more recent run (part of the same differential expression project). Supplementary data to go with David Eccles' poster about nanopore cDNA sequencing with and without mitochondrial DNA. Reference mouse mtDNA and mtDNA reads from 4T1 cells (from MinION cDNA sequencing, and from SRA run SRR6747859). fwd_4T1_BC06.correctedReads.uniqueOnly.fasta.gz -- Canu-corrected cDNA reads from Wildtype 4T1 strain, filtered as transcript-forward orientation rev_4T1_BC06.correctedReads.uniqueOnly.fasta.gz -- Canu-corrected cDNA reads from Wildtype 4T1 strain, filtered as transcript-reverse orientation fwd_4T1_BC07.correctedReads.uniqueOnly.fasta.gz -- Canu-corrected cDNA reads from 4T1ρ0 strain (no mitochondrial DNA), filtered as transcript-forward orientation rev_4T1_BC07.correctedReads.uniqueOnly.fasta.gz -- Canu-corrected cDNA reads from 4T1ρ0 strain (no mitochondrial DNA), filtered as transcript-reverse orientation Other intermediate data files are also included, see `4way_mapping_process.txt` for more details. The mpileup differential coverage script can be found here.