iu-merge-pairs error

[nikolay12]

Here is my case. I got my V1-V3 data sequenced by an external provider. They say they used the Illumina Casava pipeline version 1.8.3.

As a start I tried to generate a config file. I followed the steps listed at https://github.com/meren/illumina-utils:

I first generated a tab file listing all the sample names and the corresponding paired end fastq files. Than I ran iu-gen-configs and I was surprised that instead of generating a single config file it generated a config file for each sample.

Than I decided to merge the paired end fastq files for each sample by using iu-merge-pairs using the --compute-qual-dicts option. When I ran it for the first sample it produced the following error:

Error: Your input FASTQ files do not seem to be generated by CASAVA 1.8. Please use --ignore-deflines parameter.

I added the parameter as requested. Than I got another error message:

$ iu-merge-pairs --compute-qual-dicts --ignore-deflines 16001_posD09_CCTAAGACACTGCATA.ini
Traceback (most recent call last):
  File "/usr/local/bin/iu-merge-pairs", line 770, in <module>
    sys.exit(merger.run())
  File "/usr/local/bin/iu-merge-pairs", line 398, in run
    tile_number = self.input_1.entry.tile_number
  File "/Library/Python/2.7/site-packages/IlluminaUtils/lib/fastqlib.py", line 82, in __getattr__
    return getattr(self, '_'.join(['process', key]))()
(...)
  File "/Library/Python/2.7/site-packages/IlluminaUtils/lib/fastqlib.py", line 82, in __getattr__
    return getattr(self, '_'.join(['process', key]))()
  File "/Library/Python/2.7/site-packages/IlluminaUtils/lib/fastqlib.py", line 73, in __getattr__
    if key in ['__str__']: 
RuntimeError: maximum recursion depth exceeded in cmp

I don't know what to do now. Can you, perhaps, advise?

[meren]

Hi,

Can you please provide some example files? The first 1,000 lines of R1 and R2 reads for one of your samples in that dataset would be the best (i.e., example-R1.fastq and example-R2.fastq).

Thanks,

[nikolay12]

Thanks for your quick reply. I'm attaching the first 1000 lines. 16001_posD09_CCTAAGACACTGCATA_R1_1000.fastq.gz 16001_posD09_CCTAAGACACTGCATA_R2_1000.fastq.gz

[meren]

Hi,

I found what causes the error and I will add a control when I have a chance for the next version. Clearly it should be illegal to use --compute-qual-dicts flag when --ignore-deflines is used :( So your short-term solution is to not use --compute-qual-dicts. I am sorry for that.

The reason you are forced to use --ignore-deflines with these files is because their headers are not what we expect to see with CASAVA 1.8+. This is how it should look like:

@D4ZHLFP1:36:C10H4ACXX:8:2203:21201:39665 1:N:0:CCAT

and this is how yours look like:

@HWI-M04481:31:000000000-ARVP8:1:1101:7941:1899/1

If you look at the specification on the Illumina page, you will realize that the format they describe matches to the first one:

http://support.illumina.com/help/SequencingAnalysisWorkflow/Content/Vault/Informatics/Sequencing_Analysis/CASAVA/swSEQ_mCA_FASTQFiles.htm

Best,