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  • Problems using Bustard basecalling

    I am trying to run manual basecalling on a sample with high initial cluster intensity, but the intensity suddenly drops off. I heard using that changing the martix-cycles option.

    I ran the following command:
    /path to bustard/bustard.py / /path to sequence run/Data/Intensities --CIF --matrix-cycles=6 --maxcycle=57 --with-seq --make

    and make once again in the bustard folder created.

    I run into an error 4 issue where a .dif2 file cannot be found.

    Code:
    Controls reported in /location to seq run/Data/Intensities/Bustard1.9.4_18-04-2013_administrator/Temp/s_1_1101_controls.csv
    callBases: /location to software/OLB-1.9.4/c++/basecalling/CycleReader.cpp: ga::basecalling::BaseCycleReader::BaseCycleReader(const std::vector<boost::filesystem::basic_path<std::basic_string<char, std::char_traits<char>, std::allocator<char> >, boost::filesystem::path_traits>, std::allocator<boost::filesystem::basic_path<std::basic_string<char, std::char_traits<char>, std::allocator<char> >, boost::filesystem::path_traits> > >&, bool): line 52: 2013-Apr-18 19:18:49: No such file or directory: File /location to seq run/Data/Intensities/Bustard1.9.4_18-04-2013_administrator/L001/C1.1/s_1_1101.dif2 does not exist
    make: *** [Temp/s_1_1101.flag] Error 4

    I was wondering what I was doing wrong

    Thanks

  • #2
    What sequencer type has been used? GAIIx,HiSeq,MiSeq ..?

    CIF files are available? Quite a few people don't store CIFs due to storage capacity problems.

    When does this error occur (after make file parsing, which takes a while)?

    Can you put the complete make output somewhere? Often there is an error occurring earlier in the make process ..

    Did bustard.py spit out any error/warning?

    Btw., you probably won't need '--with-seq'. You will convert from BCL to fastq afterwards I think ..

    Comment


    • #3
      HiSeq was used. The CIF files are still available.
      I think this error occurs after make file parsing.
      The only warning from earlier is "not enough clusters to estimate phasing". I will re-run the command to get the entire output. So far, I only took a screenshot of the error..I should have a text output by later today.

      Last edited by navd; 04-19-2013, 08:07 AM.

      Comment


      • #4
        I hope you are already talking with Illumina tech support about this. If not, then you should.

        For something like this posting screen shots/including bits of log files may not help us get you the solution since the actual error may be located somewhere else in the log file.

        What version of CASAVA are you using BTW?

        Comment

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