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tk2 / RetroSeq

RetroSeq is a bioinformatics tool that searches for mobile element insertions from aligned reads in a BAM file and a library of reference transposable elements. Please read the wiki page (link below) for usage instructions. Also, there is a page on the wiki describing how the 1000 genomes CEU trio was carried out with the files and parameters used for the various steps.
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Uninitalized value error post PE parsing #4

Open biobenkj opened 8 years ago

biobenkj commented 8 years ago

When I run RetroSeq in the align mode, it gets to PE alignment parsing and then breaks. Not sure what the error means (uninitialized value before assignment?).

Input: perl bin/retroseq.pl -discover -bam ../bwa/Mtbcosmid.sorted.bam -eref ../bwa/retroseqTNlib.tab -output ../bwa/Mtbcosmidtest.candidates.tab -align

Output: RetroSeq: A tool for discovery and genotyping of transposable elements from short read alignments

Version: 1.41 Author: Thomas Keane (thomas.keane@sanger.ac.uk)

Reading -eref file: ../bwa/retroseqTNlib.tab

Min anchor quality: 20 Min percent identity: 80 Min length for hit: 36

Opening BAM (../bwa/Mtbcosmid.sorted.bam) and getting initial set of candidate mates.... Reading chromosome: pRD12F9 1075 candidate reads remain to be found after first pass.... Reading chromosome: pRD12F9 Parsing PE alignments.... Use of uninitialized value $lastLine in string ne at bin/retroseq.pl line 587. Alignment did not complete correctly

Any insight you could provide would be great!

tk2 commented 8 years ago

I suspect that your exonerate alignment did not complete. This line is where it checks the exonerate output for "-- completed exonerate analysis" from exonerate to say it completed fully. Did it maybe run over a time limit or memory on your machine?

kenza12 commented 8 years ago

Hi. I have the same problem. If someone could find a solution, it would be great to share it please. I run the tool only on one chromosome to test it. So, there is no problem of memory or time limit. Thanks

biobenkj commented 8 years ago

So the way around this @tk2 and @kenza12 is to download the latest version of exonerate (https://www.ebi.ac.uk/about/vertebrate-genomics/software/exonerate) [2.4.0], recompile and execute retroseq. Must be some issue with 2.2.0...

RoseString commented 8 years ago

I am using exonerate 2.4, but still ran into the same problem. I think there is a memory issue associated with the usage of --bestn. I have more than 100Gb memory..Not sure how to solve this.

ghost commented 7 years ago

I tried two different versions of exonerate (2.4 and 2.22) with the samples used in the tutorial. I get the same error with both versions. Below is the tail end of my output.

" Reading chromosome: GL000225.1 Reading chromosome: GL000192.1 Reading chromosome: NC_007605 Reading chromosome: hs37d5 Using reference TE locations to assign discordant mates... Screening for hits to: Alu Screening for hits to: L1HS Use of uninitialized value $lastLine in string ne at retroseq.pl line 509. Alignment did not complete correctly Parsing PE alignments.... "

I used the tutorial commands with updated paths to my files.

Is this issue going to be fixed?

dwesche commented 6 years ago

Hi @tk2 I'm getting the same error with both exonerate 2.2.0 and 2.4.0:

... 649922 candidate reads remain to be found after first pass.... Reading chromosome: chr1 ... Parsing PE alignments.... Use of uninitialized value $lastLine in string ne at /home/newmanlab/dwesche/programs/RetroSeq/bin/retroseq.pl line 509. Alignment did not complete correctly

Here's the run command: retroseq.pl -discover -align -bam /my/bam/file.bam -eref /my/eref/file.txt -output ./outfile.txt

Are there any new insights on this? Thanks!

wangruohan111 commented 5 years ago

I also have this problem and my exonerate is 2.4.0. Anyone has a solution?

tk2 commented 5 years ago

Hi - I just re-ran the NA12878 data from the wiki page and it completes just fine. The underlying cause is usually that exonerate ran out of memory, if you were running on a compute farm can you check if the process hit the memory limit?

I'm happy to have a look at specific examples if you can provide me with test data.

nikitagambhir commented 4 years ago

Hi, I had the same problem and this is certainly not a memory issue. I ran RetroSeq on 55 samples and only one sample (referred to as 'bad' sample) produced this error repeatedly. Each of my samples had 16 chromosomes. When I split the bam file of the 'bad' sample into 16 bam files (one file per chromosome) and then ran the analysis, RetroSeq worked.