Rsubread stopping half-way through align
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@campbell1933-11092
Last seen 5.1 years ago

Hi

I've been using Rsubread with a range of new and old FASTQ files and it has been working fine. The latest batch seems to be having some problems and for some of the files I get the following error after it crashes

Traceback:

 1: .C("R_align_wrapper", as.integer(n), as.character(cmd), PACKAGE = "Rsubread")

 2: align(index = "reference_index", minFragLength = 35, nthreads = 30,     readfile1 = "RS-02239140_N3_DMSO_GFP_R1.fastq.gz", output_file = "RWPE.GFP.DMSO.3.BAM")

I think i saw an earlier post about 10 months ago with a similar issue but couldn't see a resolution.

I'm using Rsubread_1.22.2

Anyone got any thoughts?

thanks

 

rsubread • 1.6k views
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Your post shows a function call but doesn't show any error message. What error message did you get? What you mean by "it crashes"? 

AFAIK there are no unresolved issues with Rsubread unexpectedly stopping.

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That was the error message and then it gives the options 1-4 for how to quit out of R. By crashes then i mean it doesn't do anything else. Maybe that's not the right word?

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The output from traceback is not an error message, so you are keeping us completely in the dark as to what may have gone wrong for you.

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Could you provide your commands and also the full screen output?

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OK will do. I'm traveling so this will take a couple of days. thanks

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The samples divided and run on Hiseq2500 v3 chemistry, high output flowcell, 100SE. The FASTQ files were then catenated.

R version 3.3.0 (2016-05-03) -- "Supposedly Educational" being run on a computer cluster.

In Rsubread

>align(index="reference_index", minFragLength=35, nthreads=30, readfile1="RS-02239142_N4_DMSO_input_R1.fastq.gz", output_file="RWPE.Inp.DMSO.1.BAM")

||                       Threads : 30                                         ||

||                  Phred offset : 33                                         ||

||                     Min votes : 3 / 10                                     ||

||    Maximum allowed mismatches : 3                                          ||

||   Maximum allowed indel bases : 5                                          ||

|| # of best alignments reported : 1                                          ||

||                Unique mapping : yes                                        ||

||                                                                            ||

\\===================== http://subread.sourceforge.net/ ======================//

 

|| The input file contains base space reads.                                  ||

|| The range of Phred scores observed in the data is [2,41] 

 

and then

 

|| The range of Phred scores observed in the data is [2,41]                   ||

....it runs for a while.....

 *** caught segfault ***

address 0x7f3a5ee09fd0, cause 'invalid permissions'

 

Traceback:

 1: .C("R_align_wrapper", as.integer(n), as.character(cmd), PACKAGE = "Rsubread")

 2: align(index = "reference_index", minFragLength = 35, nthreads = 30,     readfile1 = "RS-02239142_N4_DMSO_input_R1.fastq.gz", output_file = "RWPE.Inp.DMSO.1.BAM")

 

 

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I suspect the problem might be related to the concatenation of your fastq files. Can you try to run align() on one of the fastq files before you merge them?

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Yes, that was our thought too, but the non-concatentated file also failed

 

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Could you provide us one of your fastq files in question so we can take a close look? Which reference genome did you use in your mapping?

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Wei Shi ★ 3.3k
@wei-shi-2183
Last seen 19 hours ago
Australia/Melbourne/Olivia Newton-John …

Thanks for sending through the data. But we could not reproduce the problem you encountered. Both fastq files were successfully aligned on a linux computer.

Have you checked if you have enough disk space? Could you also provide your session info?

 

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@campbell1933-11092
Last seen 5.1 years ago

 

We mapped to Hg19

What's the best way to share a fastq file with you?

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Wei Shi ★ 3.3k
@wei-shi-2183
Last seen 19 hours ago
Australia/Melbourne/Olivia Newton-John …

Could you transfer the data via CloudStor which is fast and free to academic users?

https://cloudstor.aarnet.edu.au/sender/

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@campbell1933-11092
Last seen 5.1 years ago

send you a link to a google drive folder with two fastq files in it. One worked and one didn't. There's also a txt file of the readout from Rsubread for both files.

 

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@campbell1933-11092
Last seen 5.1 years ago

Hi - did you get the link to share the folder through google drive?

 

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@campbell1933-11092
Last seen 5.1 years ago

ok 

yes, we think it's something to do with disk space too! 

Ok, i'll get session info

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@campbell1933-11092
Last seen 5.1 years ago

So, to date, we've been doing as a fisbatch job and then tried modifying the memory allocation

fisbatch --nodes=1 --ntasks-per-node=16 --constraint=CPU-E5-2660

But still seem to be running into the same issue.  

I'm guessing fisbatch won't work always (because of memory allocation issue) and so it will have to  be a slurm job?

 

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I think you'd better seek help from your system admin.
 

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ok - thanks - yes it sounds like a system admin issue

 

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@campbell1933-11092
Last seen 5.1 years ago

But, one final question - which version of R were you using?

tx

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R 3.3.0 - the same version as you use.

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@campbell1933-11092
Last seen 5.1 years ago

thanks.

 

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