DOI: 10.5281/zenodo.7860849
title: Dataset for Multiplex-PCR detection and Nanopore-based genotyping of fish pathogens

description: This zip file contains scripts, initial fastq files, assembled genomes (public and from this study) as well as bioinformatics intermediate files used for this study. File Structure and Descriptions ├── 01_process.sh : primer trimming, length-based filtering, read alignment, alignment filtering (unique hit) and extraction of uniquely hit reads for consensus generation<br> ├── 02_consensus.sh : [need artic conda env] Generation of consensus based on uniquely-mapped reads and minimal read depth of 20x required to call a variant (or it will be masked)<br> ├── 03_cleanup.sh: General folder and intermediate file re-organization<br> ├── 04_filter.sh: [need quast conda env] statistic of consensus generated and filtering of consensus with one or more ambiguous base (N), not suitable for haplotype<br> ├── 05_cluster.sh: clustering of consensus based on 100% identity threshold to generate putative haplotype<br> ├── Amplicon_FastQ folder: uniquelly mapped fastq files for consensus generation<br> ├── BAM: alignment files generated from minimap2 used as input for the artic pipeline to identify variants<br> ├── Cluster_Rep.txt: Consensus sequences that were chosen to represent each haplotype<br> ├── Consensus folder: consensus fasta files generated for each sample containing sequences for each specific pathogen<br> ├── Coverage folder: coverage and base-level read depth for each sample and each pathogen reference genes<br> ├── Filter: individiual fasta sequences (only 1 sequence per file) for each pathogen and each sample without any ambiguous base for subsequent clustering analysis<br> ├── Full_Haplotype.fasta: all possible haplotype sequences generated for each pathogen<br> ├── Gap_Analysis.tsv: Table with percentage of gap (0-100%) for each consensus sequence generated (used for filtering)<br> ├── Haplotype folder: Intermediate file and sample-level haplotype used to infer final haplotype and generate haplotype summary<br> ├── Haplotype_summary.tsv: Table with sample ID and their respectively pathogen haplotype<br> ├── Minimap2_PAF: Intermediate alignment generated from minimap2 used to generate the count table<br> ├── Original: fastq with original naming prior to renaming based on sampleID. a script (rename.sh) was included to show renaming scheme<br> ├── primer.fasta: Primer sequences used for identifying and trimming reads with flanking primer sequence<br> ├── primer.fasta.fai: the index file for primer.fasta<br> ├── PrimerTrim folder: Primer-trimmed reads<br> ├── quast_results: consensus statistics generated by quast<br> ├── RawCount.tsv: Count table generated that can used as a input to generate figure<br> ├── RawFastq folder: Raw reads that have been renamed to reflect sample information<br> ├── readme.md: The current readme file<br> ├── ref_full_latest.fasta: Reference sequence of (gene segments) 4 pathogens e.g. TilV, ISKNV, SAG (S├── 01_process.sh : primer trimming, length-based filtering, read alignment, alignment filtering (unique hit) and extraction of uniquely hit reads for consensus generation<br> ├── 02_consensus.sh : [need artic conda env] Generation of consensus based on uniquely-mapped reads and minimal read depth of 20x required to call a variant (or it will be masked)<br> ├── 03_cleanup.sh: General folder and intermediate file re-organization<br> ├── 04_filter.sh: [need quast conda env] statistic of consensus generated and filtering of consensus with one or more ambiguous base (N), not suitable for haplotype<br> ├── 05_cluster.sh: clustering of consensus based on 100% identity threshold to generate putative haplotype<br> ├── Amplicon_FastQ folder: uniquelly mapped fastq files for consensus generation<br> ├── BAM: alignment files generated from minimap2 used as input for the artic pipeline to identify variants<br> ├── Cluster_Rep.txt: Consensus sequences that were chosen to represent each haplotype<br> ├── Consensus folder: consensus fasta files generated for each sample containing sequences for each specific pathogen<br> ├── Coverage folder: coverage and base-level read depth for each sample and each pathogen reference genes<br> ├── Filter: individiual fasta sequences (only 1 sequence per file) for each pathogen and each sample without any ambiguous base for subsequent clustering analysis <br> ├── Full_Haplotype.fasta: all possible haplotype sequences generated for each pathogen <br> ├── Gap_Analysis.tsv: Table with percentage of gap (0-100%) for each consensus sequence generated (used for filtering) <br> ├── Haplotype folder: Intermediate file and sample-level haplotype used to infer final haplotype and generate haplotype summary<br> ├── Haplotype_summary.tsv: Table with sample ID and their respectively pathogen haplotype<br> ├── Minimap2_PAF: Intermediate alignment generated from minimap2 used to generate the count table<br> ├── Original: fastq with original naming prior to renaming based on sampleID. a script (rename.sh) was included to show renaming scheme<br> ├── primer.fasta: Primer sequences used for identifying and trimming reads with flanking primer sequence<br> ├── primer.fasta.fai: the index file for primer.fasta<br> ├── PrimerTrim folder: Primer-trimmed reads<br> ├── quast_results: consensus statistics generated by quast<br> ├── RawCount.tsv: Count table generated that can used as a input to generate figure<br> ├── RawFastq folder: Raw reads that have been renamed to reflect sample information<br> ├── readme.md: The current readme file<br> ├── ref_full_latest.fasta: Reference sequence of (gene segments) 4 pathogens e.g. TilV, ISKNV, SAG (Streptococcus agalactiae), FNO (Francisella noatunensis subsp. orientalis) <br> ├── ref_full_latest.primer.fasta: Same as above but with their primer binding sequence trimmed similar to the processed reads<br> ├── ref_full_latest.primer.fasta.fai<br> ├── RenameHaplotype: Script to perform reorganization of cdhit output<br> ├── Seq.stat.tsv: Sequencing statistics<br> └── VCF: VCF files from medaka variant calling used to generate the final consensus<br> treptococcus agalactiae), FNO (Francisella noatunensis subsp. orientalis)<br> ├── ref_full_latest.primer.fasta: Same as above but with their primer binding sequence trimmed similar to the processed reads<br> ├── ref_full_latest.primer.fasta.fai<br> ├── RenameHaplotype: Script to perform reorganization of cdhit output<br> ├── Seq.stat.tsv: Sequencing statistics<br> └── VCF: VCF files from medaka variant calling used to generate the final consensus