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PAP Functional Workflow v2020.09

Annotation: Functional Phage annotation workflow, please see the guide at https://cpt.tamu.edu/training-material/topics/phage-annotation-pipeline/ for help

Step	Annotation
Step 1: Input dataset Annotations and Sequence select at runtime
Step 2: Input dataset Apollo Organism JSON File select at runtime
Step 3: Split GFF3+Fasta into separate parts Combined GFF3/Fasta File Output dataset 'output' from step 1
Step 4: Retrieve JBrowse Organism Common Name Source Autodetect from Apollo JSON Apollo Organism File Output dataset 'output' from step 2
Step 5: Remove Description Fasta file Output dataset 'fasta_out' from step 3
Step 6: Reformat GFF3 File GFF3 Annotations Output dataset 'gff_out' from step 3
Step 7: Get open reading frames (ORFs) or coding sequences (CDSs) Sequence file (nucleotides) Output dataset 'out' from step 5 Genetic code 11. Bacterial Look for ORFs or CDSs Look for ORFs (check for stop codons only, ignore start codons) Minimum length ORF/CDS (in amino acids, e.g. 30 aa = 90 bp plus any stop codon) 30 Strand to search Search both the forward and reverse strand
Step 8: NCBI BLAST+ blastn (CPT Latest) Nucleotide query sequence(s) Output dataset 'out' from step 5 Subject database/sequences Locally installed BLAST database Nucleotide BLAST database nt_current None Empty. None Empty. Type of BLAST dc-megablast - Discontiguous megablast used to find more distant (e.g., interspecies) sequences Set expectation value cutoff 0.001 Output format Tabular (extended 25 columns) Filter by TaxIDs (Can be multiple, separated by space. Will automatically include sub-IDs) 12333 1714270 10656 10472 10659 10841 10860 28883 11989 10877 79205 2 Advanced Options Show Advanced Options Filter out low complexity regions (with DUST) True Query strand(s) to search against database/subject Both Maximum hits to consider/show 0 Maximum number of HSPs (alignments) to keep for any single query-subject pair Not available. Percent identity cutoff 0.0 Word size for wordfinder algorithm Not available. Perform ungapped alignment only? False Should the query and subject defline(s) be parsed? False Restrict search of database to a given set of ID's No restriction, search the entire database Minimum query coverage per hsp (percentage, 0 to 100) 0.0 Multiple hits window size: use 0 to specify 1-hit algorithm, leave blank for default Not available. Cost to open a gap Not available. Cost to extend a gap Not available.
Step 9: GFF3 Feature Sequence Export Reference Genome From History Source FASTA Sequence Output dataset 'out' from step 5 GFF3 Annotations Output dataset 'default' from step 6 Filter for specific feature types CDS Remove description (use if blasting) False
Step 10: GFF3 Add Gene to CDS GFF3 Annotations Output dataset 'out_gff3_file' from step 7
Step 11: Blast Tabular Dice Filter Blast Results Output dataset 'output1' from step 8 Minimum dice score 0.2
Step 12: Start Codon Statistics Fasta file Output dataset 'default' from step 9
Step 13: Stop Codon Statistics Fasta file Output dataset 'default' from step 9
Step 14: Fasta Translate Fasta file Output dataset 'default' from step 9 Target output Protein Translation table to use [11] The Bacterial, Archaeal and Plant Plastid Code Remove stop characters True Convert first residue to Met True
Step 15: GFF3 Filter: Require Phage Start Reference Genome From History Source FASTA Sequence Output dataset 'out' from step 5 GFF3 Annotations Output dataset 'output' from step 10
Step 16: BlastN Results to GFF3 Choose BlastN File Format Blast 25 Column Table" Blast 25 Column Table Output dataset 'output' from step 11
Step 17: TMHMM (GFF3) Protein Fasta file Output dataset 'default' from step 14
Step 18: LipoP Input Fasta File Output dataset 'default' from step 14 HTML Output False
Step 19: Interproscan functional predictions of ORFs Fasta File Output dataset 'default' from step 14 Switch on lookup of corresponding GO annotation True Switch on lookup of correspoding InterPro annotation True Switch on lookup of correspoding Pathway annotation True Applications to run TIGRFAM: protein families based on Hidden Markov Models or HMMs PIRSF: non-overlapping clustering of UniProtKB sequences into a hierarchical order (evolutionary relationships) Panther: Protein ANalysis THrough Evolutionary Relationships SMART: identification and analysis of domain architectures based on Hidden Markov Models or HMMs PROSITE Profiles: protein domains, families and functional sites as well as associated profiles to identify them PROSITE Pattern: protein domains, families and functional sites as well as associated patterns to identify them HAMAP: High-quality Automated Annotation of Microbial Proteomes PfamA: protein families, each represented by multiple sequence alignments and hidden Markov models PRINTS: group of conserved motifs (fingerprints) used to characterise a protein family SUPERFAMILY: database of structural and functional annotation Coils: Prediction of Coiled Coil Regions in Proteins Gene3d: Structural assignment for whole genes and genomes using the CATH domain structure database SignalP Gram Positive Bacteria SignalP Gram Negative Bacteria Phobius: combined transmembrane topology and signal peptide predictor
Step 20: NCBI BLAST+ blastp (CPT Latest) Protein query sequence(s) Output dataset 'default' from step 14 Subject database/sequences Locally installed BLAST database Protein BLAST database nr None Empty. None Empty. Type of BLAST blastp - Traditional BLASTP to compare a protein query to a protein database Set expectation value cutoff 0.001 Output format BLAST XML Filter by TaxIDs (Can be multiple, separated by space. Will automatically include sub-IDs) 12333 1714270 10656 10472 10659 10841 10860 28883 11989 10877 79205 Advanced Options Hide Advanced Options
Step 21: NCBI BLAST+ blastp (CPT Latest) Protein query sequence(s) Output dataset 'default' from step 14 Subject database/sequences Locally installed BLAST database Protein BLAST database canonical_2021 None Empty. None Empty. Type of BLAST blastp - Traditional BLASTP to compare a protein query to a protein database Set expectation value cutoff 0.001 Output format BLAST XML Filter by TaxIDs (Can be multiple, separated by space. Will automatically include sub-IDs) Empty. Advanced Options Hide Advanced Options
Step 22: NCBI BLAST+ blastp (CPT Latest) Protein query sequence(s) Output dataset 'default' from step 14 Subject database/sequences Locally installed BLAST database Protein BLAST database sprot None Empty. None Empty. Type of BLAST blastp - Traditional BLASTP to compare a protein query to a protein database Set expectation value cutoff 0.001 Output format BLAST XML Filter by TaxIDs (Can be multiple, separated by space. Will automatically include sub-IDs) Empty. Advanced Options Hide Advanced Options
Step 23: Gff3 Filter: Require SD Reference Genome From History Source FASTA Sequence Output dataset 'out' from step 5 GFF3 Annotations Output dataset 'stdout' from step 15
Step 24: Rebase Wig Analysis Results Parent GFF3 annotations Output dataset 'default' from step 6 Wig Results Output dataset 'bw_i' from step 17 Map protein translated results to original DNA data True Mapping Key ID
Step 25: Rebase GFF3 features Parent GFF3 annotations Output dataset 'default' from step 6 Child GFF3 annotations to rebase against parent Output dataset 'output' from step 17 Interpro specific modifications False Map protein translated results to original DNA data True Mapping Key ID
Step 26: Rebase Wig Analysis Results Parent GFF3 annotations Output dataset 'default' from step 6 Wig Results Output dataset 'bw_o' from step 17 Map protein translated results to original DNA data True Mapping Key ID
Step 27: Rebase Wig Analysis Results Parent GFF3 annotations Output dataset 'default' from step 6 Wig Results Output dataset 'bw_m' from step 17 Map protein translated results to original DNA data True Mapping Key ID
Step 28: LipoP to GFF3 LipoP Output Output dataset 'html_file' from step 18 GFF3 for Rebasing Output dataset 'default' from step 6 Output to jBrowse True Filter for only SPII Cleavage sites True
Step 29: Rebase GFF3 features Parent GFF3 annotations Output dataset 'default' from step 6 Child GFF3 annotations to rebase against parent Output dataset 'output_gff3' from step 19 Interpro specific modifications True Map protein translated results to original DNA data True Mapping Key ID
Step 30: Interrupted gene detection tool GFF3 Annotations Output dataset 'default' from step 6 Blastp Results Output dataset 'output1' from step 20 Separation Minimum 10 Separation Maximum 2000 Percent Identity Threshold 0.3
Step 31: BlastP Results to GFF3 Choose BlastP File Format BlastXML BlastXML Input Output dataset 'output1' from step 20 Include protein alignment in Blast view in Apollo/JBrowse True
Step 32: BlastP Results to GFF3 Choose BlastP File Format BlastXML BlastXML Input Output dataset 'output1' from step 21 Include protein alignment in Blast view in Apollo/JBrowse True
Step 33: BlastP Results to GFF3 Choose BlastP File Format BlastXML BlastXML Input Output dataset 'output1' from step 22 Include protein alignment in Blast view in Apollo/JBrowse True
Step 34: Identify Lipoboxes Naive orf calls Output dataset 'stdout' from step 23 Genome Output dataset 'out' from step 5 Minimum distance in codons to start of lipobox 10 Maximum distance in codons to start of lipobox 60
Step 35: GFF3 Feature Sequence Export Reference Genome From History Source FASTA Sequence Output dataset 'out' from step 5 GFF3 Annotations Output dataset 'stdout' from step 23 Filter for specific feature types CDS Remove description (use if blasting) True
Step 36: Wig to BigWig Wig File Output dataset 'output' from step 24 Fasta File Output dataset 'out' from step 5
Step 37: Wig to BigWig Wig File Output dataset 'output' from step 26 Fasta File Output dataset 'out' from step 5
Step 38: Wig to BigWig Wig File Output dataset 'output' from step 27 Fasta File Output dataset 'out' from step 5
Step 39: Remove Annotation Feature GFF3 Annotations Output dataset 'stdout' from step 28 Remove 'remark' features True Remove 'region' features True
Step 40: Rebase GFF3 features Parent GFF3 annotations Output dataset 'default' from step 6 Child GFF3 annotations to rebase against parent Output dataset 'output' from step 31 Interpro specific modifications False Map protein translated results to original DNA data True Mapping Key ID
Step 41: Rebase GFF3 features Parent GFF3 annotations Output dataset 'default' from step 6 Child GFF3 annotations to rebase against parent Output dataset 'output' from step 32 Interpro specific modifications False Map protein translated results to original DNA data True Mapping Key ID
Step 42: Rebase GFF3 features Parent GFF3 annotations Output dataset 'default' from step 6 Child GFF3 annotations to rebase against parent Output dataset 'output' from step 33 Interpro specific modifications False Map protein translated results to original DNA data True Mapping Key ID
Step 43: Fasta Translate Fasta file Output dataset 'default' from step 35 Target output Protein Translation table to use [11] The Bacterial, Archaeal and Plant Plastid Code Remove stop characters True Convert first residue to Met True
Step 44: LipoP Input Fasta File Output dataset 'default' from step 43 HTML Output False
Step 45: NCBI BLAST+ blastp (CPT Latest) Protein query sequence(s) Output dataset 'default' from step 43 Subject database/sequences Locally installed BLAST database Protein BLAST database spanindbv2 None Empty. None Empty. Type of BLAST blastp - Traditional BLASTP to compare a protein query to a protein database Set expectation value cutoff 0.001 Output format BLAST XML Filter by TaxIDs (Can be multiple, separated by space. Will automatically include sub-IDs) Empty. Advanced Options Hide Advanced Options
Step 46: TMHMM (GFF3) Protein Fasta file Output dataset 'default' from step 43
Step 47: LipoP to GFF3 LipoP Output Output dataset 'html_file' from step 44 GFF3 for Rebasing Output dataset 'stdout' from step 23 Output to jBrowse True Filter for only SPII Cleavage sites True
Step 48: Rebase GFF3 features Parent GFF3 annotations Output dataset 'stdout' from step 23 Child GFF3 annotations to rebase against parent Output dataset 'output' from step 46 Interpro specific modifications False Map protein translated results to original DNA data True Mapping Key ID
Step 49: Intersect and Adjacent GFF3 Annotations A Output dataset 'default' from step 48 GFF3 Annotations B Output dataset 'default' from step 34 Adjacency Window Size 50 Only allow adjacencies for features on the same strand True
Step 50: Intersect and Adjacent GFF3 Annotations A Output dataset 'default' from step 48 GFF3 Annotations B Output dataset 'stdout' from step 47 Adjacency Window Size 50 Only allow adjacencies for features on the same strand True
Step 51: Remove Annotation Feature GFF3 Annotations Output dataset 'oa' from step 49 Remove 'remark' features True Remove 'region' features True
Step 52: Remove Annotation Feature GFF3 Annotations Output dataset 'ob' from step 49 Remove 'remark' features True Remove 'region' features True
Step 53: Remove Annotation Feature GFF3 Annotations Output dataset 'ob' from step 50 Remove 'remark' features True Remove 'region' features True
Step 54: Remove Annotation Feature GFF3 Annotations Output dataset 'oa' from step 50 Remove 'remark' features True Remove 'region' features True
Step 55: JBrowse Reference genome to display Use a genome from history Select the reference genome Output dataset 'fasta_out' from step 3 Produce Standalone Instance False Genetic Code 11. The Bacterial, Archaeal and Plant Plastid Code JBrowse-in-Galaxy Action Update exising JBrowse Instance Previous JBrowse Instance Output dataset 'jbrowse' from step 4 Track Groups Track Group 1 Track Category #date# Functional Annotation / Sequence Analysis / Phage Annotation Tracks Annotation Track 1 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'output' from step 30 This is match/match_part data False Maximum size of track chunks 2000000 Index this track False JBrowse Track Type [Advanced] Canvas Features CanvasFeatures Options [Advanced]: Transcript type Empty. Subfeatures type Empty. Implied UTRs False JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label Empty. JBrowse style.description note,description JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Track Group 2 Track Category #date# Functional Annotation / Sequence Analysis / Structural Annotation Tracks Annotation Track 1 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'default' from step 25 This is match/match_part data False Maximum size of track chunks 2000000 Index this track False JBrowse Track Type [Advanced] HTML Features HTMLFeatures Options [Advanced]: Top level features Empty. JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label product,name,id JBrowse style.description note,description JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Annotation Track 2 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'default' from step 29 This is match/match_part data False Maximum size of track chunks 2000000 Index this track False JBrowse Track Type [Advanced] Canvas Features CanvasFeatures Options [Advanced]: Transcript type Empty. Subfeatures type Empty. Implied UTRs False JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label signature_desc,product,name,id JBrowse style.description note,description,name JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Annotation Track 3 Track Type BigWig XY BigWig Track Data Output dataset 'bigwig' from step 36,Output dataset 'bigwig' from step 37 Use XYPlot True Show variance band False Track Scaling Specify Min/Max Track minimum 0 Track maximum 1 Visual Scaling Linear Use MultiBigWig Plugin False JBrowse Color Options [Advanced]: Color Specification Automatically selected Bicolor Pivot Zero JBrowse Custom Track Config [Advanced]: Custom Track Config Options Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Annotation Track 4 Track Type BigWig XY BigWig Track Data Output dataset 'bigwig' from step 38 Use XYPlot True Show variance band False Track Scaling Specify Min/Max Track minimum 0 Track maximum 1 Visual Scaling Linear Use MultiBigWig Plugin False JBrowse Color Options [Advanced]: Color Specification Automatically selected Bicolor Pivot Zero JBrowse Custom Track Config [Advanced]: Custom Track Config Options Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Annotation Track 5 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'default' from step 39 This is match/match_part data False Maximum size of track chunks 2000000 Index this track False JBrowse Track Type [Advanced] Canvas Features CanvasFeatures Options [Advanced]: Transcript type Empty. Subfeatures type Empty. Implied UTRs False JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label product,name,id JBrowse style.description note,description JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Track Group 3 Track Category #date# Functional Annotation / Blast / Nucleotide Annotation Tracks Annotation Track 1 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'output' from step 16 This is match/match_part data True Match Part Feature Type match Maximum size of track chunks 2000000 Index this track False JBrowse Track Type [Advanced] Blast Features JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label product,name,id JBrowse style.description note,description JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Track Group 4 Track Category #date# Functional Annotation / Blast / Protein Annotation Tracks Annotation Track 1 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'default' from step 40,Output dataset 'default' from step 42,Output dataset 'default' from step 41 This is match/match_part data True Match Part Feature Type match Maximum size of track chunks 5000000 Index this track False JBrowse Track Type [Advanced] Blast Features JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label product,name,id JBrowse style.description note,description JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Track Group 5 Track Category #date# Functional Annotation / Sequence Analysis / Spanin Annotation Tracks Annotation Track 1 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'default' from step 52,Output dataset 'default' from step 53 This is match/match_part data False Maximum size of track chunks 2000000 Index this track False JBrowse Track Type [Advanced] Canvas Features CanvasFeatures Options [Advanced]: Transcript type Empty. Subfeatures type Empty. Implied UTRs False JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label product,name,id JBrowse style.description note,description JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Annotation Track 2 Track Type Blast XML BlastXML Track Data Output dataset 'output1' from step 45 Features used in Blast Search Output dataset 'stdout' from step 23 Minimum Gap Size 2 Is this a protein blast search? True Index this track False JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label description JBrowse style.description Hit_titles JBrowse style.height 600px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override Annotation Track 3 Track Type GFF/GFF3/BED Features GFF/GFF3/BED Track Data Output dataset 'default' from step 51,Output dataset 'default' from step 54 This is match/match_part data False Maximum size of track chunks 2000000 Index this track False JBrowse Track Type [Advanced] Canvas Features CanvasFeatures Options [Advanced]: Transcript type Empty. Subfeatures type Empty. Implied UTRs False JBrowse Styling Options [Advanced]: JBrowse style.className feature JBrowse style.label product,name,id JBrowse style.description note,description JBrowse style.height 10px JBrowse maxHeight 600 JBrowse Feature Score Scaling & Coloring Options [Advanced]: Color Score Algorithm Ignore score Color Selection Automatically selected JBrowse Custom Track Config [Advanced]: Custom Track Config Options JBrowse Contextual Menu options [Advanced]: Track Menus Track Visibility Off for new users Override Apollo Plugins No - Do not Override Override Apollo Draggability No - Do not Override General JBrowse Options [Advanced]: Default Location Empty. Track Padding 20 Enable Share Link True About the Browser Empty. Show Track List True Show Navigation True Show Overview True Show Menu True Hide Genome Options False Plugins: BlastView True Combo Track Selector False GC Content False None Empty.
Step 56: Create or Update Organism JBrowse HTML Output Output dataset 'output' from step 55 Organism Common Name Source Autodetect from Apollo JSON Apollo Organism File Output dataset 'output' from step 2 Genus Empty. Species Empty. Is Organism Public False Grant access to a user group None Remove old data directory False Update genome sequences False None no
Step 57: Annotate Apollo Organism Listing Output dataset 'output' from step 56 Force the tracklist on in JBrowse False

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PAP Functional Workflow v2020.09

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