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  • BWA - Error in Run Time

    I have installed BWA. And build index of hg19 using command:

    Code:
    bwa index -a bwtsw hg19.fa
    Now I am trying to find alignment using command:

    Code:
    $ ./bwa aln hg.fa SRR493081_1.fastq > aln_sa.sai
    [bwa_aln] 17bp reads: max_diff = 2
    [bwa_aln] 38bp reads: max_diff = 3
    [bwa_aln] 64bp reads: max_diff = 4
    [bwa_aln] 93bp reads: max_diff = 5
    [bwa_aln] 124bp reads: max_diff = 6
    [bwa_aln] 157bp reads: max_diff = 7
    [bwa_aln] 190bp reads: max_diff = 8
    [bwa_aln] 225bp reads: max_diff = 9
    [bwa_aln] fail to locate the index
    [main] Version: 0.6.2-r126
    [main] CMD: ./bwa aln hg.fa SRR493081_1.fastq
    [main] Real time: 0.000 sec; CPU: 0.010 sec
    However, gives above error. Can anybody helps me why is this giving such error.

  • #2
    According to your statement,
    this commond
    $ ./bwa aln hg.fa SRR493081_1.fastq > aln_sa.sai

    should be

    $ ./bwa aln hg19.fa SRR493081_1.fastq > aln_sa.sai

    Because hg.fa is different with hg19.fa, bwa can't find the index.

    Comment


    • #3
      I have a similar problem and cannot get bwa alignment to work.

      I completed the index successfully
      Code:
      $ bwa index -a bwtsw -p hg19 hg19.fa
      bwa_index] Pack FASTA... 27.96 sec
      [bwa_index] Construct BWT for the packed sequence...
      [BWTIncCreate] textLength=6274322528, availableWord=453484340
      [BWTIncConstructFromPacked] 10 iterations done. 100000000 characters processed.
      [BWTIncConstructFromPacked] 20 iterations done. 200000000 characters processed.
      .
      .
      .
      [BWTIncConstructFromPacked] 690 iterations done. 6264217232 characters processed.
      [bwt_gen] Finished constructing BWT in 695 iterations.
      [bwa_index] 3556.44 seconds elapse.
      [bwa_index] Update BWT... 21.30 sec
      [bwa_index] Pack forward-only FASTA... 18.66 sec
      [bwa_index] Construct SA from BWT and Occ... 1118.66 sec
      [main] Version: 0.7.5a-r405
      [main] CMD: bwa index -a bwtsw -p hg19 hg19.fa
      [main] Real time: 4768.228 sec; CPU: 4743.022 sec
      then when I attempted the alignment:

      Code:
      $ bwa aln -q 28 -t 12 hg19.fa sample1_1.fastq.gz > sample1_1.sai
      [bwa_aln] 17bp reads: max_diff = 2
      [bwa_aln] 38bp reads: max_diff = 3
      [bwa_aln] 64bp reads: max_diff = 4
      [bwa_aln] 93bp reads: max_diff = 5
      [bwa_aln] 124bp reads: max_diff = 6
      [bwa_aln] 157bp reads: max_diff = 7
      [bwa_aln] 190bp reads: max_diff = 8
      [bwa_aln] 225bp reads: max_diff = 9
      [bwa_aln] fail to locate the index
      Even when I use the absolute path i get :
      Code:
      $ bwa aln -q 28 -t 12 /NGSsoftware/hg19.fa 8-A182_1.fastq.gz > 8-A182_1.sai
      [bwa_aln] 17bp reads: max_diff = 2
      [bwa_aln] 38bp reads: max_diff = 3
      [bwa_aln] 64bp reads: max_diff = 4
      [bwa_aln] 93bp reads: max_diff = 5
      [bwa_aln] 124bp reads: max_diff = 6
      [bwa_aln] 157bp reads: max_diff = 7
      [bwa_aln] 190bp reads: max_diff = 8
      [bwa_aln] 225bp reads: max_diff = 9
      [bwa_aln] fail to locate the index
      I was wondering if there were permission problems but even after changing permissions of to all users rwx it still will not work.

      Any suggestions? I am using Mac OSX 10.8.4 with Xcode installed.

      Comment


      • #4
        Since you used "-p hg19" as the prefix for the output database you need to include that as the prefix for the database.

        Try the following:

        Code:
        $ bwa aln -q 28 -t 12 /NGSsoftware/hg19 8-A182_1.fastq.gz > 8-A182_1.sai

        Comment


        • #5
          Originally posted by GenoMax View Post
          Since you used "-p hg19" as the prefix for the output database you need to include that as the prefix for the database.

          Try the following:

          Code:
          $ bwa aln -q 28 -t 12 /NGSsoftware/hg19 8-A182_1.fastq.gz > 8-A182_1.sai
          Thank you GenoMax, this worked perfectly until I tried to perform bwa sampe:

          Code:
          $ bwa sampe –r '@RG\tID:sample8\tLB:sample8\tSM:sample8\tPL:ILLUMINA' /NGSsoftware/hg19  8-A182_1.sai  8-A182_2.sai 8-A182_1.fastq.gz 8-A182_2.fastq.gz 8-A182.sam
          [bwa_sai2sam_pe] fail to locate the index
          If I remove the -r argument, bwa sampe will run, but I wish to have the read groups added.

          Comment


          • #6
            Can you post a listing of the files in the /NGSsoftware directory?

            Code:
            $ ls -l /NGSsoftware/hg19*

            Comment


            • #7
              Here are the files

              Code:
              $ ls -l /NGSsoftware/hg19*
              -rw-r--r--  1 kwilliams  wheel        6693  5 Sep 13:38 /NGSsoftware/hg19.amb
              -rw-r--r--  1 kwilliams  wheel        3635  5 Sep 13:38 /NGSsoftware/hg19.ann
              -rw-r--r--  1 kwilliams  wheel  3101804844  5 Sep 13:38 /NGSsoftware/hg19.bwt
              -rw-r--r--  1 kwilliams  wheel  3163842112  5 Sep 08:06 /NGSsoftware/hg19.fa
              -rw-r--r--  1 kwilliams  wheel   775451187  5 Sep 13:38 /NGSsoftware/hg19.pac
              -rw-r--r--  1 kwilliams  wheel  1550902424  5 Sep 13:57 /NGSsoftware/hg19.sa

              Comment


              • #8
                Hi, just wondering if anyone has insight into the problem above regarding read groups and bwa?
                Many thanks.

                Comment


                • #9
                  Have you tried to run the command with double quotes (") instead of single?

                  Code:
                  $ bwa sampe –r "@RG\tID:sample8\tLB:sample8\tSM:sample8\tPL:ILLUMINA" /NGSsoftware/hg19  8-A182_1.sai  8-A182_2.sai 8-A182_1.fastq.gz 8-A182_2.fastq.gz 8-A182.sam
                  It is confusing that you chose to use "hg19" as your prefix name for the indexes.
                  Last edited by GenoMax; 09-15-2013, 02:07 PM.

                  Comment


                  • #10
                    Yes, that also comes up with
                    Code:
                    [bwa_sai2sam_pe] fail to locate the index

                    Comment


                    • #11
                      Should have asked this before. Are you using a Mac? Did you compile bwa yourself?

                      Comment


                      • #12
                        Yes I am using a mac. I have Xcode installed and compiled bwa using Homebrew.

                        Comment


                        • #13
                          There were some problems with bwa on a Mac in past (http://seqanswers.com/forums/showthread.php?t=24133). I am not sure if those persist/are still relevant. Perhaps someone else who is currently running the latest bwa (0.7.5) on Mac can confirm.

                          What version of OS X (and Xcode) are you running?

                          Comment

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