mirDeep2 mapper.pl is making an empty reads_collapsed_vs_genome.arf file - help?!

[jmccreig]

I'm having trouble running mirDeep2, and I was hoping someone could help me out. I ran mirDeep2 fine the other day, but now it's mysteriously no longer working. The first step of using bowtie to process the reference genome seems to be working fine, but my issue is when I go to the next step:

perl mapper.pl hg19miRNA_reads.fasta -c -j -l 18 -m -p HUMAN_Whole_Genome -s HUMAN_reads_collapsed.fa -t HUMAN_reads_collapsed_vs_genome.arf -v

I get the following errors:

discarding sequences with non-canonical letters
discarding short reads
collapsing reads
mapping reads to genome index
trimming unmapped nts in the 3' ends
Mapping statistics

#desc total mapped unmapped %mapped %unmapped
Use of uninitialized value $count2 in subtraction (-) at mapper.pl line 630.
total: 293297 Use of uninitialized value $count2 in print at mapper.pl line 630.
293297 Use of uninitialized value $count2 in division (/) at mapper.pl line 631.
Use of uninitialized value $count2 in division (/) at mapper.pl line 631.
0.000 1.000
Use of uninitialized value in subtraction (-) at mapper.pl line 633.
seq: 293297 Use of uninitialized value in print at mapper.pl line 633.
293297 Use of uninitialized value in division (/) at mapper.pl line 634.
Use of uninitialized value in division (/) at mapper.pl line 634.
0.000 1.000

I'm getting these divide by zero errors because there are absolutely no reads being mapped to the genome - my reads_collapsed_vs_genome.arf file that's generated is completely empty.

When I ran this the other day using the same set of data, it worked perfectly (generated pdfs and everything). In the interrum, I installed Biopieces in order to use their find_adaptor and clip_adaptor functions, because I only had about 25% of my reads mapping, and I thought some of my reads may have sequencing into the adaptors and that was affecting my ability to map them back to the genome. But since installing that, I keep getting this mirDeep2 error - nothing is mapping! Even if I go back and use all the data that worked before, it doesn't work.

Any ideas? Thanks so much for helping.

[jmccreig]

I figured it out, and wanted to share the simple solution since I had seen this problem posted elsewhere unanswered... Turns out I just didn't have enough RAM, and our cluster was killing my job halfway through the initial bowtie step, which was ruining all downstream steps. Once I allocated more memory to my job, everything worked perfectly.

[okapick]

Me too . Have you solved? Have you asked to the authors?



/home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl out_WGS_k55.fasta -c -i -j -p ty55_1 -s ty55_1h.fa -t ty55_1h_vs_genome.arf
touch: no se puede efectuar `touch' sobre «mapper.log_bak»: Permiso denegado
Reads file contained a pattern with more than 1024 sequence characters.
Please truncate reads and quality values and and re-run Bowtie.
Offending read: ENTRY330941
terminate called after throwing an instance of 'int'

#desc total mapped unmapped %mapped %unmapped
Use of uninitialized value $count2 in subtraction (-) at /home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl line 630, <IN> line 111967.
Use of uninitialized value $count in subtraction (-) at /home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl line 630, <IN> line 111967.
total: Use of uninitialized value $count in print at /home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl line 630, <IN> line 111967.
Use of uninitialized value $count2 in print at /home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl line 630, <IN> line 111967.
0 Use of uninitialized value $count in division (/) at /home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl line 631, <IN> line 111967.
Use of uninitialized value $count2 in division (/) at /home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl line 631, <IN> line 111967.
Illegal division by zero at /home/soba/Documentos/Ensambladores/mirdeep2/mapper.pl line 631, <IN> line 111967.
┌─( soba ) ─ ( ~/Documentos/Ensambladores/mirdeep2/tutorial_dir ) ───────────── (13:09:52 - mar jun 17)

[BioRyder]

Hi,

The reads_collapsed_vs_genome.arf file empty problem is happening may be due to bowtie versions.Please check the bowtie version which you installed in your machine and bowtie which you have used for indexing.If both are different (like version 1,1.01,1.1 etc) ,then reads_collapsed_vs_genome.arf will be empty.