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Old 07-29-2014, 06:19 AM   #1
wintergreen36
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Default bcl2fastq run error

[email protected]:~$ /usr/local/bin/configureBclToFastq.pl --input-dir /media/vanessa/UUI/ngs/Data/Intensities/BaseCalls --output-dir /media/vanessa/UUI Unaligned --sample-sheet /media/vanessa/UUI/ngs/Data/Intensities/BaseCalls/SampleSheet.csv
"my" variable $value masks earlier declaration in same statement at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 760.
syntax error at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 747, near "$variable qw(ELAND_FASTQ_FILES_PER_PROCESS)"
Global symbol "$variable" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 749.
syntax error at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 751, near "$directory qw(ELAND_GENOME)"
Global symbol "$self" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 753.
Global symbol "$directory" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 753.
Global symbol "$project" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 753.
Global symbol "$sample" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 753.
Global symbol "$lane" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 753.
Global symbol "$barcode" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 753.
Global symbol "$reference" requires explicit package name at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 753.
syntax error at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm line 761, near "}"
/usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment/Config.pm has too many errors.
Compilation failed in require at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment.pm line 61.
BEGIN failed--compilation aborted at /usr/local/lib/bcl2fastq-1.8.3/perl/Casava/Alignment.pm line 61.
Compilation failed in require at /usr/local/bin/configureBclToFastq.pl line 239.
BEGIN failed--compilation aborted at /usr/local/bin/configureBclToFastq.pl line 239.

can any body solve this errors
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Old 07-29-2014, 06:23 AM   #2
GenoMax
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Is there a space in the --output-dir declaration "/vanessa/UUI Unaligned"?
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Old 07-29-2014, 11:58 PM   #3
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Default @ Genomax

Yes ther is space ,

[email protected]:~$ /usr/local/bin/configureBclToFastq.pl --input-dir /media/vanessa/UUI/ngs/Data/Intensities/BaseCalls --output-dir /media/vanessa/UUI/ Unaligned --sample-sheet /media/vanessa/UUI/ngs/Data/Intensities/BaseCalls/SampleSheet.csv

Please watch the screen shot of usr/local/bin

is there any other requirements other than existed ,like simple XML executables , to begin ?
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Old 07-30-2014, 03:01 AM   #4
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When you specify an --output-dir location the "Unaligned" directory is automatically created by the configureBclToFastq.pl script in that directory. You do not need to specify it on the command line.

Omit the "Unaligned" from your command line and the program should work.

You may want to consider adding "--fastq-cluster-count 0" (without quotes) to your command line if you want the sequence output in a single file as opposed to 4 millon fastq sequences each in more than one file.
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Old 07-30-2014, 04:41 AM   #5
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Default solved

I solved it , Thanks buddy
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Old 08-05-2014, 12:20 AM   #6
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@ Genomax

Thanks for the above guidelines
Finally I could able to run the Bcl2fastq , i tried to demultiplex 2X100 pe nextra exome data with 8 bp indices ,unfortunately all fastq files were moved to undetermined_Indices folder , in which sample_lane1 , sample_lane2 has no data except sample sheet ,rest of the folders lane 3 to lane 8 goy sample sheet + fasq.gz files

the command I uses is /usr/local/bin/configureBclToFastq.pl --input-dir
<BaseCalls_dir> --output-dir <Unaligned> --sample-sheet
<BaseCalls_dir>/SampleSheet.csv --use-bases-mask y100n,I8n,Y100N

Last edited by wintergreen36; 08-05-2014 at 12:21 AM. Reason: missing content
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Old 08-05-2014, 03:05 AM   #7
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If lanes 3-8 worked fine then the format of your sample sheet as well as the command line is fine. Did you look at the Demultiplex_Stats.htm file to ascertain that the demultiplexing had worked as expected (i.e. more reads assigned to samples than are in the "undetermined" pool)? Is the situation reverse for lanes 1 and2 (i.e more reads in undetermined row for lanes 1/2)?

Start looking at the fastq ID headers of the "lane1_undetermined*" files to see if there are any problems with the tag reads (i.e. N's etc).
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Old 08-11-2014, 09:09 PM   #8
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Hello GEnomax thanks for ur support I SUCCESFULY INSTALLED bcl2fastq BUt while running a project i got the follwing error ,

[2014-08-12 13:23:41] [vanessa] [Temp/L006_R1_demux_summary.xml] Processing BCL files for lane 6, tile 2316, read 1
[2014-08-12 13:24:14] [vanessa] [Temp/L006_R1_demux_summary.xml] runtime error: boost::filesystem::rename: No such file or directory: "/media/vanessa/Sandor_1/Unaligned/Undetermined_indices/Sample_lane6/lane6_Undetermined_L006_R1_001.fastq.tmp.gz", "/media/vanessa/Sandor_1/Unaligned/Undetermined_indices/Sample_lane6/lane6_Undetermined_L006_R1_001.fastq.gz"
make: *** [Temp/L006_R1_demux_summary.xml] Error 2

Could you please guide me how to over come this error

thanks
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Old 08-12-2014, 03:05 AM   #9
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Unfortunately not enough information (at least for me) in that error snippet. Have you looked further up in the log file to see if there was any other errors (missing BCL/stat files etc)? Were there any notes from the facility that ran the flowcell about lane 6 experiencing problems during run (no clusters may be a possibility if the error above is real)?
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Old 08-12-2014, 04:05 AM   #10
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I ran this there was no errors during the run
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Old 08-12-2014, 04:22 AM   #11
GenoMax
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Have you tried removing entries for lane 6 samples from your samplesheet and checking to see if the remaining lanes demultiplex?
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Old 08-12-2014, 04:31 AM   #12
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I haven't , There was no boost libs installed in my system , I just updated libboost all ; started run again
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Old 08-12-2014, 04:32 AM   #13
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What was the impact of these boost lib on Bcl2fastq run?
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Old 08-12-2014, 04:35 AM   #14
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There was no boost lib in my system , what's the impact of boost lib on bcl2fastq run ? I just installed lib boost and started the bcl2fastq run again.
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Old 08-12-2014, 04:43 AM   #15
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That is odd. You have been able to run bcl2fastq before and I would have though that Boost libraries are required for the software to function. Let us see what happens with the new run.
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