De-Multiplexing with Je-Demultiplex-Illu

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1

I just tried to use Je-Demultiplex-Illu to demultiplex my Illumina reads. I thought I entered everything correctly, but I got the following error. Any suggestions how to fix this?

Picked up _JAVA_OPTIONS: -Xmx4G -Xms256m
Exception in thread “main” htsjdk.samtools.SAMException: Sequence header must start with @: ��BC*�}�n�J���|��a��/�,d��lo�ܕ��[&�K�rV#��n7}`RL��|�?C�e�7 at line 1 in fastq /data/dnb10/galaxy_db/files/5/3/5/dataset_5357985d-c1a8-42d2-b6e1-2ace4621f4cd.dat
at htsjdk.samtools.fastq.FastqReader.readNextRecord(FastqReader.java:98)
at htsjdk.samtools.fastq.FastqReader.(FastqReader.java:64)
at htsjdk.samtools.fastq.FastqReader.(FastqReader.java:52)
at org.embl.gbcs.je.jemultiplexer.BarcodeValidator.getRealReadLength(BarcodeValidator.java:765)
at org.embl.gbcs.je.jemultiplexer.BarcodeValidator.checkBarcodesLengthWithIndexFiles(BarcodeValidator.java:746)
at org.embl.gbcs.je.jemultiplexer.JemultiplexerIllumina.validateBarcodes(JemultiplexerIllumina.java:567)
at org.embl.gbcs.je.jemultiplexer.AbstractJemultiplexer.customCommonCommandLineValidation(AbstractJemultiplexer.java:618)
at org.embl.gbcs.je.jemultiplexer.JemultiplexerIllumina.customCommandLineValidation(JemultiplexerIllumina.java:190)
at picard.cmdline.CommandLineProgram.parseArgs(CommandLineProgram.java:267)
at picard.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:151)
at picard.cmdline.CommandLineProgram.instanceMainWithExit(CommandLineProgram.java:147)
at org.embl.gbcs.je.Je.main(Je.java:112)

2

This message from the tool is indicating that the data is not in a fastq format it can understand.

That could be the compression state being wrong with the datatype, or something is actually wrong with one or more files inside of them (truncated?).

Try to Upload using all defaults to let Galaxy guess the datatype format. Then you can run a tool like Fastq info to check the technical parts. If that all checks out, then you can proceed to other tools.

Hope this helps! We have lots of related Q&A here at the forum. I added some tags to your topic that should help, too. :slight_smile: