featurecounts not working on mirbase annotation file
0
0
Entering edit mode
13 months ago

Hello I am trying to analyze miRNA-seq data but I am having problems with the mapping. I always get pretty much 0 counts with the built in annotation file, so I got one from miRBase. However, I always get an error when I use it, it doesn't even run. I thought the error could have been in the structure of the file, so I downloaded one from GENCODEand compared both structures. They didn't seem different, but the one from GENCODE runs just like the built-in one (runs but gives zero counts). So I have no idea what is going on.

Here are the gff3 structures:

miRBase:

chr1    .   miRNA_primary_transcript    12425986    12426106    .   +   .   ID=MI0021869;Alias=MI0021869;Name=mmu-mir-6341          
chr1    .   miRNA   12426016    12426038    .   +   .   ID=MIMAT0025084;Alias=MIMAT0025084;Name=mmu-miR-6341;Derives_from=MI0021869         
chr1    .   miRNA_primary_transcript    20679010    20679082    .   +   .   ID=MI0000249;Alias=MI0000249;Name=mmu-mir-206           
chr1    .   miRNA   20679017    20679039    .   +   .   ID=MIMAT0017004;Alias=MIMAT0017004;Name=mmu-miR-206-5p;Derives_from=MI0000249           

GENCODE:

chr1    HAVANA  gene    3073253 3074322 .   +   .   ID=ENSMUSG00000102693.1;gene_id=ENSMUSG00000102693.1;gene_type=TEC;gene_status=KNOWN;gene_name=4933401J01Rik;level=2;havana_gene=OTTMUSG00000049935.1           
chr1    HAVANA  transcript  3073253 3074322 .   +   .   ID=ENSMUST00000193812.1;Parent=ENSMUSG00000102693.1;gene_id=ENSMUSG00000102693.1;transcript_id=ENSMUST00000193812.1;gene_type=TEC;gene_status=KNOWN;gene_name=4933401J01Rik;transcript_type=TEC;transcript_status=KNOWN;transcript_name=4933401J01Rik-001;level=2;transcript_support_level=NA;tag=basic;havana_gene=OTTMUSG00000049935.1;havana_transcript=OTTMUST00000127109.1         
chr1    HAVANA  exon    3073253 3074322 .   +   .   ID=exon:ENSMUST00000193812.1:1;Parent=ENSMUST00000193812.1;gene_id=ENSMUSG00000102693.1;transcript_id=ENSMUST00000193812.1;gene_type=TEC;gene_status=KNOWN;gene_name=4933401J01Rik;transcript_type=TEC;transcript_status=KNOWN;transcript_name=4933401J01Rik-001;exon_number=1;exon_id=ENSMUSE00001343744.1;level=2;transcript_support_level=NA;tag=basic;havana_gene=OTTMUSG00000049935.1;havana_transcript=OTTMUST00000127109.1           
chr1    ENSEMBL gene    3102016 3102125 .   +   .   ID=ENSMUSG00000064842.1;gene_id=ENSMUSG00000064842.1;gene_type=snRNA;gene_status=KNOWN;gene_name=Gm26206;level=3            
chr1    ENSEMBL transcript  3102016 3102125 .   +   .   ID=ENSMUST00000082908.1;Parent=ENSMUSG00000064842.1;gene_id=ENSMUSG00000064842.1;transcript_id=ENSMUST00000082908.1;gene_type=snRNA;gene_status=KNOWN;gene_name=Gm26206;transcript_type=snRNA;transcript_status=KNOWN;transcript_name=Gm26206-201;level=3;transcript_support_level=NA;tag=basic         

any idea of what could be wrong?

usegalaxy miRNA-seq • 658 views
ADD COMMENT
0
Entering edit mode

What does your featureCounts command line look like?

ADD REPLY
0
Entering edit mode

on the one that works. I change nothing when I use the one from mirBASE.

export FC_PATH=$(command -v featureCounts | sed 's@/bin/featureCounts$@@') &&  featureCounts  -a ${FC_PATH}/annotation/mm10_RefSeq_exon.txt -F "SAF"  -o "output" -T ${GALAXY_SLOTS:-2}  -s  0  -Q  0               --minOverlap  1 --fracOverlap 0 --fracOverlapFeature 0     '/data/dnb08/galaxy_db/files/1/2/6/dataset_12675960-3c58-4a48-aba6-f4ca750e7eff.dat'  && grep -v "^#" "output" | sed -e 's|/data/dnb08/galaxy_db/files/1/2/6/dataset_12675960-3c58-4a48-aba6-f4ca750e7eff.dat|HISAT2 on data 384: aligned reads (BAM)|g' > body.txt && cut -f 1,7 body.txt > '/data/jwd02f/main/060/487/60487873/outputs/galaxy_dataset_39d1a976-03e0-48b6-9aab-2dfcee0947d6.dat'    && sed -e 's|/data/dnb08/galaxy_db/files/1/2/6/dataset_12675960-3c58-4a48-aba6-f4ca750e7eff.dat|HISAT2 on data 384: aligned reads (BAM)|g' 'output.summary' > '/data/jwd02f/main/060/487/60487873/outputs/galaxy_dataset_199b296d-de69-44ef-82a6-3866f9fa65ee.dat'
ADD REPLY

Login before adding your answer.

Traffic: 4434 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6