Mikado commands

Mikado documentation

1 Run mikado configure to generate the configuration and scoring files

cd /home/train/Annotation_workshop/Mikado/Chr3-1065466-1464870/6_Full_Mikado

mikado configure --full --list list.txt --reference Inputs/Assembly/Athaliana_447_TAIR10.Chr3.fa --junctions Inputs/Junctions/portcullis.pass.junctions.regionA.bed -bt Inputs/Homology/selected_species.fa --scoring plant.yaml --copy-scoring plant.yaml configuration.toml

2 Run mikado prepare to remove redundancy, fix or remove problematic transcripts and extract the transcript sequences

mikado prepare -p 1 --json-conf configuration.toml

3 Run Prodigal to call ORFs

prodigal -g 1 -f gff -i mikado_prepared.fasta -o mikado_prepared.cds.gff

4 Run diamond for homology scores

diamond blastx --threads 1 -q mikado_prepared.fasta -d Homology/selected_species -o mikado_prepared_matches.tsv --outfmt 6 qseqid sseqid pident length mismatch gapopen qstart qend sstart send evalue bitscore ppos btop

5 Run mikado serialise to load ORFs, homology hits and junctions to the database

mikado serialise -p 1 --tsv mikado_prepared_matches.tsv --orfs mikado_prepared.cds.gff --json-conf configuration.toml

6 Run mikado pick to identify gene loci and select the best transcripts

mikado pick -p 1 --subloci-out mikado.subloci.gff3 --monoloci-out mikado.monoloci.gff3 --output-dir MyOutput --json-conf configuration.toml

7 Run mikado compare to compare our selected mikado pick transcripts to the reference genemodels

mikado compare -r Athaliana_447_Araport11.gene_exons.regionA.gtf -p MyOutput/mikado.loci.gff3

8 Run mikado util stats to generate the gene stats for the mikado genemodels and the reference genemodels

mikado util stats MyOutput/mikado.loci.gff3 mikado.loci.gff3.stats

mikado util stats Athaliana_447_Araport11.gene_exons.regionA.gtf Athaliana_447_Araport11.gene_exons.regionA.gtf.stats

9 Side by side comparison of the stats

paste <(cut -f1,2,3 Athaliana_447_Araport11.gene_exons.regionA.gtf.stats |sed  "1i Reference\t") <(cut -f2,3 mikado.loci.gff3.stats |sed  "1i Mikado") |tabulate -1 -s '\t' -f grid |tee Reference_and_Mikado.stats

10 Try editing the scoring or configuration file OR using one of the alternative configuration and scoring files

mikado pick -p 1 --subloci-out mikado.subloci.gff3 --monoloci-out mikado.monoloci.gff3 --output-dir MyOutput2 --json-conf Alt_configs/reat_transcriptome_plants_configuration.toml --scoring-file Alt_configs/reat_transcriptome_plants_scoring.yaml

Does this improve the gene models?

The entire process can be run using the provided bash script commands.sh # you will be prompted to provide alternative configuration and scoring files, press enter for the indicated default. Try rerunning with the alternative configuration and scoring files under Alt_configs/ and comparing the mikado compare stats outputs