/scripts/STARlongMatchAnnot_commands_ENCODE_denovo_param.txt
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SEEK ID: https://fairdomhub.org/data_files/3184?version=1
Filename: STARlongMatchAnnot_commands_ENCODE_denovo_param.txt
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# MPE

#potato transcriptome v2 (evigene) STARlong mapping of transcripts to reference genome and MatchAnnot to get transcript annotations
#last edited: 16.05.2019

cd /DATA/markop/potato_tr_v2_matchannot 
#mkdir star_index
mkdir STARlong.ENCODE.denovo

#ulimit default 1024
ulimit -n 10000

#create genome index for DM v4.04 using the merged iTag+PGSC gtf
# STARlong \
# --runThreadN 28 \
# --runMode genomeGenerate \
# --genomeDir ./star_index \
# --genomeFastaFiles ./potatoDM404genome/StPGSC4.04n_2018-01-18.fasta \
# --sjdbGTFfile ./potatoDM404genome/StPGSC4.04n_PGSC-ITAG-merged_representative-transcript_genes_2019-04-23.gtf \
# --limitGenomeGenerateRAM 210000000000

#mapping Rywal transcriptome
STARlong --genomeLoad LoadAndExit --genomeDir ./star_index

#parameters from STARmanual ENCODE options, SAM attributes copied from Liz 
#more parameters here: https://groups.google.com/forum/#!topic/rna-star/E2eevlGWFbQ
# and here: https://github.com/PacificBiosciences/IsoSeq_SA3nUP/wiki/Old-Tutorial:-Optimizing-STAR-aligner-for-Iso-Seq-data
# also added multimap chimera detection 
# Between the two RH haplotypes, 6.6 Mb of sequence could be aligned with 96.5% identity, corresponding to 1 SNP per 29?bp and 1 indel per 253?bp (average length 10.4?bp)

STARlong \
--runMode alignReads \
--outSAMattributes NH HI NM MD \
--outFilterType BySJout \
--outFilterMultimapNmax 20 \
--alignSJoverhangMin 8 \
--alignSJDBoverhangMin 1 \
--outFilterMismatchNmax 999 \
--outFilterMismatchNoverReadLmax 0.08\
--alignIntronMin 20 \
--alignIntronMax 1000000 \
--alignMatesGapMax 1000000 \
--outSAMtype SAM \
--limitBAMsortRAM 100000000000 \
--runThreadN 28 \
--genomeDir ./star_index \
--readFilesIn ../../_A_01_evigene/intermediate_tr2aacds_Rywal/okayset/all.okay.tr \
--outFileNamePrefix ./STARlong.ENCODE.denovo/Rywal.tr.okay_DM_STARlong. \
--seedPerReadNmax 100000 \
--seedPerWindowNmax 1000 \
--seedSearchLmax 30 \
--seedSearchStartLmax 30 \
--alignTranscriptsPerReadNmax 100000 \
--alignTranscriptsPerWindowNmax 10000 \
--scoreGapNoncan -20 \
--scoreDelOpen -1 \
--scoreDelBase -1 \
--scoreInsOpen -1 \
--scoreInsBase -1 \
--chimMultimapNmax 20 \
--chimSegmentMin 100


STARlong \
--runMode alignReads \
--outSAMattributes NH HI NM MD \
--outFilterType BySJout \
--outFilterMultimapNmax 20 \
--alignSJoverhangMin 8 \
--alignSJDBoverhangMin 1 \
--outFilterMismatchNmax 999 \
--outFilterMismatchNoverReadLmax 0.08\
--alignIntronMin 20 \
--alignIntronMax 1000000 \
--alignMatesGapMax 1000000 \
--outSAMtype SAM \
--limitBAMsortRAM 100000000000 \
--runThreadN 28 \
--genomeDir ./star_index \
--readFilesIn ../../_A_01_evigene/intermediate_tr2aacds_Rywal/okayset/all.okalt.tr \
--outFileNamePrefix ./STARlong.ENCODE.denovo/Rywal.tr.okalt_DM_STARlong. \
--seedPerReadNmax 100000 \
--seedPerWindowNmax 1000 \
--seedSearchLmax 30 \
--seedSearchStartLmax 30 \
--alignTranscriptsPerReadNmax 100000 \
--alignTranscriptsPerWindowNmax 10000 \
--scoreGapNoncan -20 \
--scoreDelOpen -1 \
--scoreDelBase -1 \
--scoreInsOpen -1 \
--scoreInsBase -1 \
--chimMultimapNmax 20 \
--chimSegmentMin 100


#mapping PW363 transcriptome
STARlong \
--runMode alignReads \
--outSAMattributes NH HI NM MD \
--outFilterType BySJout \
--outFilterMultimapNmax 20 \
--alignSJoverhangMin 8 \
--alignSJDBoverhangMin 1 \
--outFilterMismatchNmax 999 \
--outFilterMismatchNoverReadLmax 0.08\
--alignIntronMin 20 \
--alignIntronMax 1000000 \
--alignMatesGapMax 1000000 \
--outSAMtype SAM \
--limitBAMsortRAM 100000000000 \
--runThreadN 28 \
--genomeDir ./star_index \
--readFilesIn ../../_A_01_evigene/intermediate_tr2aacds_PW363/okayset/all.okay.tr \
--outFileNamePrefix ./STARlong.ENCODE.denovo/PW363.tr.okay_DM_STARlong. \
--seedPerReadNmax 100000 \
--seedPerWindowNmax 1000 \
--seedSearchLmax 30 \
--seedSearchStartLmax 30 \
--alignTranscriptsPerReadNmax 100000 \
--alignTranscriptsPerWindowNmax 10000 \
--scoreGapNoncan -20 \
--scoreDelOpen -1 \
--scoreDelBase -1 \
--scoreInsOpen -1 \
--scoreInsBase -1 \
--chimMultimapNmax 20 \
--chimSegmentMin 100



STARlong \
--runMode alignReads \
--outSAMattributes NH HI NM MD \
--outFilterType BySJout \
--outFilterMultimapNmax 20 \
--alignSJoverhangMin 8 \
--alignSJDBoverhangMin 1 \
--outFilterMismatchNmax 999 \
--outFilterMismatchNoverReadLmax 0.08\
--alignIntronMin 20 \
--alignIntronMax 1000000 \
--alignMatesGapMax 1000000 \
--outSAMtype SAM \
--limitBAMsortRAM 100000000000 \
--runThreadN 28 \
--genomeDir ./star_index \
--readFilesIn ../../_A_01_evigene/intermediate_tr2aacds_PW363/okayset/all.okalt.tr \
--outFileNamePrefix ./STARlong.ENCODE.denovo/PW363.tr.okalt_DM_STARlong. \
--seedPerReadNmax 100000 \
--seedPerWindowNmax 1000 \
--seedSearchLmax 30 \
--seedSearchStartLmax 30 \
--alignTranscriptsPerReadNmax 100000 \
--alignTranscriptsPerWindowNmax 10000 \
--scoreGapNoncan -20 \
--scoreDelOpen -1 \
--scoreDelBase -1 \
--scoreInsOpen -1 \
--scoreInsBase -1 \
--chimMultimapNmax 20 \
--chimSegmentMin 100




#mapping Desiree transcriptome
STARlong \
--runMode alignReads \
--outSAMattributes NH HI NM MD \
--outFilterType BySJout \
--outFilterMultimapNmax 20 \
--alignSJoverhangMin 8 \
--alignSJDBoverhangMin 1 \
--outFilterMismatchNmax 999 \
--outFilterMismatchNoverReadLmax 0.08\
--alignIntronMin 20 \
--alignIntronMax 1000000 \
--alignMatesGapMax 1000000 \
--outSAMtype SAM \
--limitBAMsortRAM 100000000000 \
--runThreadN 28 \
--genomeDir ./star_index \
--readFilesIn ../../_A_01_evigene/intermediate_tr2aacds_Desiree/okayset/all.okay.tr \
--outFileNamePrefix ./STARlong.ENCODE.denovo/Desiree.tr.okay_DM_STARlong. \
--seedPerReadNmax 100000 \
--seedPerWindowNmax 1000 \
--seedSearchLmax 30 \
--seedSearchStartLmax 30 \
--alignTranscriptsPerReadNmax 100000 \
--alignTranscriptsPerWindowNmax 10000 \
--scoreGapNoncan -20 \
--scoreDelOpen -1 \
--scoreDelBase -1 \
--scoreInsOpen -1 \
--scoreInsBase -1 \
--chimMultimapNmax 20 \
--chimSegmentMin 100




STARlong \
--runMode alignReads \
--outSAMattributes NH HI NM MD \
--outFilterType BySJout \
--outFilterMultimapNmax 20 \
--alignSJoverhangMin 8 \
--alignSJDBoverhangMin 1 \
--outFilterMismatchNmax 999 \
--outFilterMismatchNoverReadLmax 0.08\
--alignIntronMin 20 \
--alignIntronMax 1000000 \
--alignMatesGapMax 1000000 \
--outSAMtype SAM \
--limitBAMsortRAM 100000000000 \
--runThreadN 28 \
--genomeDir ./star_index \
--readFilesIn ../../_A_01_evigene/intermediate_tr2aacds_Desiree/okayset/all.okalt.tr \
--outFileNamePrefix ./STARlong.ENCODE.denovo/Desiree.tr.okalt_DM_STARlong. \
--seedPerReadNmax 100000 \
--seedPerWindowNmax 1000 \
--seedSearchLmax 30 \
--seedSearchStartLmax 30 \
--alignTranscriptsPerReadNmax 100000 \
--alignTranscriptsPerWindowNmax 10000 \
--scoreGapNoncan -20 \
--scoreDelOpen -1 \
--scoreDelBase -1 \
--scoreInsOpen -1 \
--scoreInsBase -1 \
--chimMultimapNmax 20 \
--chimSegmentMin 100




STARlong --genomeLoad Remove --genomeDir ./star_index


#sorting SAM files (same as done by Liz)
for i in $(ls ./STARlong.ENCODE.denovo/*.sam | sed s/.sam// )
do sort -k 3,3 -k 4,4n ${i}.sam > ${i}.sorted.sam
done

#MatchAnnot
#/home/administrator/Software/MatchAnnot/matchAnnot.py --gtf=./potatoDM404genome/StPGSC4.04n_PGSC-ITAG-merged_representative-transcript_genes_2019-04-23.gtf --format=alt\
   ./STARlong.ENCODE.denovo/Desiree.tr.okalt_DM_STARlong.Aligned.out.sorted.sam > ./STARlong.ENCODE.denovo/Desiree.tr.okalt_DM_STARlong.Aligned.out.sorted.sam.matchAnnot.txt

for i in $(ls ./STARlong.ENCODE.denovo/*.sorted.sam )
do
   /home/administrator/Software/MatchAnnot/matchAnnot.py --gtf=./potatoDM404genome/StPGSC4.04n_PGSC-ITAG-merged_representative-transcript_genes_2019-04-23.gtf --format=alt\
   ${i} > ${i}.matchAnnot.txt
done


#parsing the matchannot results: EvigenetrID  DMgeneID DMtrID exon_match match_score fwd/rev
# grep "result:" Desiree.tr.okay_DM_STARlong.Aligned.out.sorted.sam.matchAnnot.txt | tr -s ' ' | sed 's/5-3:/fwd/g' | awk -F'[ ]' '{print $2, $3, $4, $6, $8, $9}' > Desiree.tr.okay_DM_STARlong.Aligned.out.sorted.sam.matchAnnot.parsed.txt
# actually I don't need info on fwd/rev

for i in $(ls ./STARlong.ENCODE.denovo/*.sorted.sam.matchAnnot.txt | sed s/.txt// )
  do grep "result:" ${i}.txt | tr -s ' ' | awk -F'[ ]' '{print $2, $3, $4, $6, $8}' > ${i}.parsed.txt
done
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