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  • GMAP gsnap input error

    Hi everyone,

    I'm trying to run gsnap programm with paired-end Illumina data. This is the command:

    gsnap -D /data/database -d hg19 s_5.1.fastq .s_5.2.fastq -A sam >> s_5__hg19.sam

    After two hours of running and 1,000,000 lines in the resulting sam file I get the following error:

    ========================
    Signal received: Segmentation fault
    Problem sequence: DD7DT8Q1:4:1101:20988:58254#ACACAA (100 bp)
    >DD7DT8Q1:4:1101:20988:58254#ACACAA
    GATGAAGGGAGGGATAGAGGGAGAATGGATGAAGGGAGAGTGGATGGATGGATGGGTGTAGGGTGGATGTGTGGAGGGCAGGTCGGAAGAGCACACGCCT
    TCCATCCATACATGCACCCTTCATCCATCCATTCATCCACTCTCCCTTCATCCATTCTCCCTCTATCCCTCCCTTCATCAGATCGGAAGAGCGTCGTGTA
    /usr/local/gridengine/default/spool/makoshark/job_scripts/225677: line 12: 30062 Aborted (core dumped)
    ========================

    This sequence looks like this:
    ========================
    @DD7DT8Q1:4:1101:20988:58254#ACACAA/1
    GATGAAGGGAGGGATAGAGGGAGAATGGATGAAGGGAGAGTGGATGGATGGATGGGTGTAGGGTGGATGTGTGGAGGGCAGGTCGGAAGAGCACACGCCT
    +DD7DT8Q1:4:1101:20988:58254#ACACAA/1
    c\_b`[ddd`ddd`_`dXddZQcccf^^TeYTTV[cT]`^`a`_BBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBBB
    ========================

    I don't see the problem with this sequence, but anyway gsnap doesn't accept it. Does anyone know what the problem might be?

    Thanks in advance,
    Irina

  • #2
    Developer of GMAP and GSNAP

    Hi Irina,

    I have just created an account on SEQanswers to help answer questions about GMAP and GSNAP (and GSTRUCT when it is finally released).

    To answer your issue, there is a known bug in version 2011-09-14 for reads of 100 bp or longer. This bug has been fixed in the latest release, 2011-10-01, available at the distribution Web site at http://research-pub.gene.com/gmap. Sorry for any inconvenience. If you have any further issues, you can send email directly to me at [email protected].

    Regards,

    Thomas D. Wu, M.D., Ph.D.
    Senior Scientist
    Bioinformatics and Computational Biology
    Genentech, Inc.
    South San Francisco, CA 94080, USA
    Last edited by twu; 10-03-2011, 12:59 PM.

    Comment


    • #3
      Hi Thomas,

      I am getting similar error as Irina with the current version 2012-07-20 (http://research-pub.gene.com/gmap/sr...2-07-20.tar.gz)

      thanks
      Uma

      Comment


      • #4
        Error

        Hi Uma,

        Please feel free to send more details to me at [email protected], and I can take a closer look. I have had no other bug reports from others so far on the 2012-07-20 release, so I would be interested to know what you are seeing.

        Regards,

        Tom

        Comment


        • #5
          Error

          I am seeing a similar error with version 2012-11-09

          Comment


          • #6
            I am trying to align paired end reads with GSNAP (version 2013-03-27) but I am getting a similar error.

            My input fastq file is in the format described in the readme:

            =================================
            >@HWI-ST273:279:C05D8ACXX:7:1101:12952:5966
            CTTTTAAGAGGATTCTTTCTGAGCAAATTTCAGTTATGAGGTAGATTCTAATTGTTCAATGAGTAACATTCTCAGGATTTCTAACTCAAGTGATCAGATC
            GATCACTTGAGTTAGAAATCCTGAGAATGTTACTCATTGAACAATTAGAATCTACCTCATAACTGAAATTTGCTCAGAAAGAATCCTCTTAAAAGAGATC
            +
            CCCFFFFFHHHHHJJJJJJJJGIJJJJJJJJJJIJJJJJJJFHIJJJJJJIJJJIJJJJJJJJIJJIJJJJJJJJJJJJIJJJJJHHHHHEEFFFFEFEC
            CCCFFFFFHHHHHJJJJJJJJJJJJJJJJJJJJJJJJJIJJJJJJJJJJJIJJJJJJJJJJJJJJJJJJJJJJJJIJJJJJJHHHHHFFFFFFFEDEEED
            >@HWI-ST273:279:C05D8ACXX:7:1101:19991:5793
            CCCGACTCCTTCAACCCACCAATGCCAAGCTGAGCCCTGCTTTCTCCCACAAGTCCCATATCTGGTGCCAAAAGTATTATTCCCAAGCTTGATCCATAAC
            GGCCTTCCAAGGTGAAACCTGAGTTATGGATCAAGCTTGGGAATAATACTTTTGGCACCAGATATGGGACTTGTGGGAGAAAGCAGGGCTCAGCTTGGCA
            +
            CCCFFFFFHHHHHJJJJJJJJIJJJJJJJJJJJJJJJJJJJJJJJJJJJJIJJJJJJJJJJJJHHHHHFFFEDE6;CEEEFEEDDCDDDDDDDDCDCCD@
            CCCFFFFFHHHHFHJJJJJIJHIHIIIJIHIJJIIJJGIJJHIJJJIJJJGIIJJJJJJJJIJIIJJJIIHHHHHFFFFDEEECDDDDDDDDDDDDDDD@
            ============================

            etc.

            I called GSNAP as follows:
            gsnap -t 4 -A sam -m 4 -d mm9 -s mm9.introns.iit -A sam CRF0003_GsnapForm.fastq > test.sam

            and got:

            290272 unique splicesites...
            290272 splicesites are valid...splicetrie_obs has 290272 entries...done
            GMAP modes: pairsearch, indel_knownsplice, terminal, improvement
            Starting alignment
            Unexpected pairtype 0
            Aborted

            It did create a partial alignment with (test.sam had about 4k lines in what looked to be good SAM format), but it seems to be quitting partway through.

            Any ideas as to what I might be doing wrong?
            Last edited by afkoeppel; 04-10-2013, 10:34 AM.

            Comment


            • #7
              Originally posted by afkoeppel View Post
              I am trying to align paired end reads with GSNAP (version 2013-03-27) but I am getting a similar error.

              My input fastq file is in the format described in the readme:

              =================================
              >@HWI-ST273:279:C05D8ACXX:7:1101:12952:5966
              CTTTTAAGAGGATTCTTTCTGAGCAAATTTCAGTTATGAGGTAGATTCTAATTGTTCAATGAGTAACATTCTCAGGATTTCTAACTCAAGTGATCAGATC
              GATCACTTGAGTTAGAAATCCTGAGAATGTTACTCATTGAACAATTAGAATCTACCTCATAACTGAAATTTGCTCAGAAAGAATCCTCTTAAAAGAGATC
              +
              CCCFFFFFHHHHHJJJJJJJJGIJJJJJJJJJJIJJJJJJJFHIJJJJJJIJJJIJJJJJJJJIJJIJJJJJJJJJJJJIJJJJJHHHHHEEFFFFEFEC
              CCCFFFFFHHHHHJJJJJJJJJJJJJJJJJJJJJJJJJIJJJJJJJJJJJIJJJJJJJJJJJJJJJJJJJJJJJJIJJJJJJHHHHHFFFFFFFEDEEED
              >@HWI-ST273:279:C05D8ACXX:7:1101:19991:5793
              CCCGACTCCTTCAACCCACCAATGCCAAGCTGAGCCCTGCTTTCTCCCACAAGTCCCATATCTGGTGCCAAAAGTATTATTCCCAAGCTTGATCCATAAC
              GGCCTTCCAAGGTGAAACCTGAGTTATGGATCAAGCTTGGGAATAATACTTTTGGCACCAGATATGGGACTTGTGGGAGAAAGCAGGGCTCAGCTTGGCA
              +
              CCCFFFFFHHHHHJJJJJJJJIJJJJJJJJJJJJJJJJJJJJJJJJJJJJIJJJJJJJJJJJJHHHHHFFFEDE6;CEEEFEEDDCDDDDDDDDCDCCD@
              CCCFFFFFHHHHFHJJJJJIJHIHIIIJIHIJJIIJJGIJJHIJJJIJJJGIIJJJJJJJJIJIIJJJIIHHHHHFFFFDEEECDDDDDDDDDDDDDDD@
              ============================

              etc.

              I called GSNAP as follows:
              gsnap -t 4 -A sam -m 4 -d mm9 -s mm9.introns.iit -A sam CRF0003_GsnapForm.fastq > test.sam

              and got:

              290272 unique splicesites...
              290272 splicesites are valid...splicetrie_obs has 290272 entries...done
              GMAP modes: pairsearch, indel_knownsplice, terminal, improvement
              Starting alignment
              Unexpected pairtype 0
              Aborted

              It did create a partial alignment with (test.sam had about 4k lines in what looked to be good SAM format), but it seems to be quitting partway through.

              Any ideas as to what I might be doing wrong?
              Quick Update: When I limited my input file to 1000 reads it worked just fine. There's something it's not liking about some (but not all) of my sequences.

              One of the offending sequences is:
              >@HWI-ST273:279:C05D8ACXX:7:1201:18203:90202
              GTATAGTAATGCCTGCGGCTAGCACTGGTAGTGATAATAGGAGCAGTACGGCTGTAATAAGTACGGATCAGACAAATAGTGGAGTTTGATACTAGATCGG
              AGTATCAAACTCCACTATTTGTCTGATCCGTACTTATTACAGCCGTACTGCTCCTATTATCACTACCAGTGCTAGCCGCAGGCATTACTATACAGATCGG
              +
              BB@FFFEDHHHHHJJJJIGJJJJIJIII?DEHGGIJJJGIJIJIGH@F@HIIGI@ECHEGIEHHEDFFDEECE=C?CDDDDADB:ACC3@ACD@>@C@BD
              CCCF<DEFHFHHGJJIJIJJJHIJJIIIIJHHIJIJJIIJJJIGGGFHIHHHHHIJJJIIIJJJJJIIJGHJIIHHHHFDDDDDDDDCDEEDDEDDDDDD

              ...but I'm not seeing any obvious difference between it and the others.

              Comment


              • #8
                Please use version 2013-03-31

                That particular bug was fixed in version 2013-03-31. Sorry for the inconvenience.

                Regards,

                Tom

                Comment


                • #9
                  Hi,

                  I still run into the segmentation fault in the 2014-05-15 version. Here is the error message.

                  =====================================
                  Looking for index files in directory /references/H.Sapiens/hg20/gmap/reference
                  Pointers file is reference.ref153offsets64meta
                  Offsets file is reference.ref153offsets64strm
                  Positions file is reference.ref153positions
                  Offsets compression type: bitpack64
                  Allocating memory for ref offset pointers, kmer 15, interval 3...done (134,217,744 bytes, 1.32 sec)
                  Allocating memory for ref offsets, kmer 15, interval 3...done (470,734,416 bytes, 4.19 sec)
                  Allocating memory for ref positions, kmer 15, interval 3...done (3,913,147,304 bytes, 34.40 sec)
                  GMAP modes: pairsearch, indel_knownsplice, terminal, improvement
                  Starting alignment
                  Signal received: Segmentation fault
                  Problem sequence: FCC2EJEACXX:8:1214:12079:6463#ATCACGAT (90 bp)
                  >FCC2EJEACXX:8:1214:12079:6463#ATCACGAT
                  ACGCATACACACAGAGACATACATACACATGTGCATACACACATACAAACACACGCATACACACATACACACGCATACACACAGAGACAT
                  TGTGTATGCATGTGTGTGTATGCATATGTGTATGTGTGTATGCGTGTGTATGTCTCTGTGTGTATGCATGTGTATGTGTATATGTGTGTG
                  /bin/sh: line 1: 41324 Aborted (core dumped) SGE_RREQ="-q all.q -pe BWA 4 -l h_vmem=8G" /usr/local/gmap/gmap-2014-05-15/bin/gsnap --format sam --nthreads 4 --db reference --novelsplicing 1 --npaths 1 --batch 4 --quiet-if-excessive --dir /references/H.Sapiens/hg20/gmap read1.qc.fq read2.qc.fq > output.hg20.sam
                  =====================================

                  I used the hg20 analysis set as the reference. The gmap index worked fine for some samples, but not this one.

                  Thanks,
                  Zuotian

                  Comment


                  • #10
                    [GSNAP] Signal received: Segmentation fault

                    I am trying to align single end reads with GSNAP (version 2014-09-22) but I am getting a similar error.

                    My input fastq file is in the format described in the readme:

                    ============[Sample 1]===============
                    @HWUSI-EAS1759R:40:FC:6:30:3714:19225 1:N:0:AGTCAA
                    CGGGGTTTCACCATCTTAACCAGGCTGGTCTCAAACTCCTGACCTCAGGTG
                    +
                    GGG4GIIIIIIHIIIIIGIIHIIIIHIIIIIIIIIIIIIIDHIIIIIIICG
                    ==================================

                    ============[Sample 2]===============
                    @HWUSI-EAS1759R:52:FC:2:20:5324:12059 1:N:0:TGACCA
                    CTCAGGTGATCCAACCGTCTCGGCCTCCCAAAGTGCTGGGATTACAGGCGT
                    +
                    IIIIHGFGGGIEIHIIGHIHIIGIIIIHIIB>EAEIFIIGHBHEGIIHIIC
                    ==================================

                    etc.

                    I called GSNAP as follows:
                    gsnap -d hg19 -s hg19.splicesites --query-unk-mismatch=1 -N 1 -B 4 -t 20 -O -A sam --force-xs-dir --force-single-end sample1.fastq >output.sam


                    and got:
                    [Sample 1]
                    Signal received: Segmentation fault
                    Problem sequence: HWUSI-EAS1759R:40:FC:6:30:3714:19225 (51 bp)
                    >HWUSI-EAS1759R:40:FC:6:30:3714:19225
                    CGGGGTTTCACCATCTTAACCAGGCTGGTCTCAAACTCCTGACCTCAGGTG

                    [Sample 2]
                    Signal received: Segmentation fault
                    Problem sequence: HWUSI-EAS1759R:52:FC:2:20:5324:12059 (51 bp)
                    >HWUSI-EAS1759R:52:FC:2:20:5324:12059
                    CTCAGGTGATCCAACCGTCTCGGCCTCCCAAAGTGCTGGGATTACAGGCGT


                    I wanna to see where is alternative splicing, or fusion gene...
                    Any ideas as to what I might be doing wrong?

                    thank you,
                    Sharon

                    Comment


                    • #11
                      solve my Segmentation fault problem

                      I think I figure out the problem....

                      My command line should "-B 2" instead of "-B 4" for single-end reads.

                      and when I change the optional....the "Segmentation fault" is disappear

                      Comment

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