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  • bam-load

    Hi Everyone ,
    I am trying to use bam-load function to convert an SRA file to bam file .



    bam-load.2.1.13 [options] <bam-file>

    Summary:
    Load a BAM formatted data file

    Example:
    bam-load.2.1.13 -o /tmp/SRZ123456 -k analysis.bam.cfg 123456.bam
    Can anyone can explain what options . What is exactly analysis.bam.cfg
    what input's it takes and what is output in detail.

    Thanks,
    Anusha.Ch

  • #2
    SRA file usually contains unmapped reads, you need to use the fastq-dump in the sra-tool kit to dump it to fastq, then you map it with bowtie which gives you a sam file, you then can use samtools to convert sam to bam.

    Comment


    • #3
      Using Bowtie to map fastq to sam file format

      Hi Everyone ,
      Can anyone give examples of using bowtie to map from fastq to sam format .

      I did try all the options it is not working for me


      /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2 -1 /raid/development/anusha/SRR203400.fastq --U - -S /raid/development/anusha/SRR203400.sam

      /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19|-U /-1 ./SRR203400.fastq -S /raid/development/anusha/SRR203400.sam

      /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19 |-U </raid/software/development/SRR203400.fastq> -S /raid/development/anusha/SRR203400.sam

      /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19| -u/ -1 /raid/development/anusha/SRR203400.fastq -S /raid/development/anusha/SRR2034001.sam

      /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19 | -u 1 /raid/development/anusha/SRR203400.fastq -S /raid/development/anusha/SRR2034001.sam

      bowtie2 [options]* -x <bt2-idx> {-1 <m1> -2 <m2> | -U <r>} [-S <sam>]
      can anyone what is option u ,what it takes with detailed example

      my errors are always like this
      Error: Must specify at least one read input with -U/-1/-2
      bowtie2-align exited with value 1

      Thanks,
      Anusha.Ch

      Comment


      • #4
        Originally posted by AnushaC View Post
        Hi Everyone ,
        Can anyone give examples of using bowtie to map from fastq to sam format .

        I did try all the options it is not working for me


        /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2 -1 /raid/development/anusha/SRR203400.fastq --U - -S /raid/development/anusha/SRR203400.sam

        /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19|-U /-1 ./SRR203400.fastq -S /raid/development/anusha/SRR203400.sam

        /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19 |-U </raid/software/development/SRR203400.fastq> -S /raid/development/anusha/SRR203400.sam

        /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19| -u/ -1 /raid/development/anusha/SRR203400.fastq -S /raid/development/anusha/SRR2034001.sam

        /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2/hg19 | -u 1 /raid/development/anusha/SRR203400.fastq -S /raid/development/anusha/SRR2034001.sam

        bowtie2 [options]* -x <bt2-idx> {-1 <m1> -2 <m2> | -U <r>} [-S <sam>]
        can anyone what is option u ,what it takes with detailed example

        my errors are always like this
        Error: Must specify at least one read input with -U/-1/-2
        bowtie2-align exited with value 1

        Thanks,
        Anusha.Ch
        Firstly, you should start a new thread next time. Secondly, the error message is because you misspecify the bowtie2 command. "{-1 <m1> -2 <m2> | -U <r>}" means you either use both -1 and -2 (i.e., you have paired-end reads) or you use only -U (i.e. you have single end reads). A proper command would be more like:
        Code:
        /raid/software/src/bowtie2 -q -x /raid/references-and-indexes/hg19/bowtie2 -U /raid/development/anusha/SRR203400.fastq -S /raid/development/anusha/SRR203400.sam

        Comment


        • #5
          Thanks for reply ,
          It is working


          Thanks,
          Anusha.Ch

          Comment


          • #6
            Hi Everyone ,
            First of all I am sorry that I am not using new thread . I want to use bowtie default 0.12.7 and i want to use option -n 3 ,- v3 and -k 2 .
            opment/anusha/SRR203400.sam
            Could not locate a Bowtie index corresponding to basename "s"
            Command: bowtie -n 3 -v 3 -k 2 --sam s /raid/references-and-indexes/hg19/bowtie-indexes /raid/development/anusha/SRR203400.fastq
            anusha@cn1:/raid/development/anusha> bowtie -n 3 -v 3 -k 2 --sam -c /raid/references-and-indexes/hg19/bowtie-indexes /raid/development/anusha/SRR203400.fastq > /raid/development/anusha/SRR203400.sam
            Could not locate a Bowtie index corresponding to basename "/raid/references-and-indexes/hg19/bowtie-indexes"
            Command: bowtie -n 3 -v 3 -k 2 --sam -c /raid/references-and-indexes/hg19/bowtie-indexes /raid/development/anusha/SRR203400.fastq
            anusha@cn1:/raid/development/anusha>

            I did not understand what is going wrong

            this is my reference index location and files

            cd /raid/references-and-indexes/hg19/bowtie-indexes
            anusha@cn1:/raid/references-and-indexes/hg19/bowtie-indexes> ls
            chrM-tRNA-rRNA-genes_c.1.ebwt chrM-tRNA-rRNA-genes_c.4.ebwt chrM-tRNA-rRNA-genes.fa hg19.3.ebwt hg19_c.2.ebwt hg19_c.rev.1.ebwt hg19.rev.2.ebwt
            chrM-tRNA-rRNA-genes_c.2.ebwt chrM-tRNA-rRNA-genes_c.rev.1.ebwt hg19.1.ebwt hg19.4.ebwt hg19_c.3.ebwt hg19_c.rev.2.ebwt
            chrM-tRNA-rRNA-genes_c.3.ebwt chrM-tRNA-rRNA-genes_c.rev.2.ebwt hg19.2.ebwt hg19_c.1.ebwt hg19_c.4.ebwt hg19.rev.1.ebwt
            anusha@cn1:/raid/references-and-indexes/hg19/bowtie-indexes>

            can anybody explain in detail how to use bowtie with clear example


            Thanks very much ,
            Anusha.Ch

            Comment


            • #7
              Code:
              bowtie -v 3 -k 2 --sam /raid/references-and-indexes/hg19/bowtie-indexes/hg19 /raid/development/anusha/SRR203400.fastq > /raid/development/anusha/SRR203400.sam
              You can't use both -n and -v, they're mutually exclusive.

              Comment


              • #8
                Loading the bam data to IGV

                Hi Everyone ,
                Can any one can explain me in detail step by step how to upload your bam data to IGV . I have my bam file which is sorted and indexed.

                Thanks In Advance ,
                Anusha.ch

                Comment


                • #9
                  If your bam file is really large its not very advisable to open it with IGV, you'd rather use Samtools to create a subset of the BAM file for the regions you need, sort and index it and open it with IGV.

                  Otherwise, its just as simple as File->Open->Point to your bam in directory, change the reference genome based on your needs from the drop down menu. If your genome is not already there, you need to import it, which is slightly complicated.

                  Comment


                  • #10
                    Opening file with IGV

                    Hi ,
                    I have my bam file sorted and indexed that is bam and bai files can anyone explain how to open those file with IGV in detail every step very clearly


                    Thanks,
                    Anusha.Ch

                    Comment


                    • #11
                      Originally posted by AnushaC View Post
                      Hi ,
                      I have my bam file sorted and indexed that is bam and bai files can anyone explain how to open those file with IGV in detail every step very clearly


                      Thanks,
                      Anusha.Ch
                      Click on "File" and then "Load", just like opening anything else. Have a read through the IGV user guide.

                      Comment


                      • #12
                        Trying making files HTMl

                        anusha@cn1:/raid/development/anusha/IGV> chmod 755 SRR203400.bowtieold.bam
                        anusha@cn1:/raid/development/anusha/IGV> chmod 755 SRR203400.sort.bam.bam
                        anusha@cn1:/raid/development/anusha/IGV> chmod 755 SRR203400.sort.bam.bam.bai
                        anusha@cn1:/raid/development/anusha/IGV> ln -s /raid/development/anusha/IGV/SRR203400.bowtieold.bam /raid/development/anusha/IGV/SRR203400.bowtieold.bam
                        ln: failed to create symbolic link ‘/raid/development/anusha/IGV/SRR203400.bowtieold.bam’: File exists
                        anusha@cn1:/raid/development/anusha/IGV> cd IGV
                        anusha@cn1:/raid/development/anusha/IGV/IGV> ls
                        IGV SRR203400.bowtieold.bam SRR203400.sort.bam.bam SRR203400.sort.bam.bam.bai
                        anusha@cn1:/raid/development/anusha/IGV/IGV> cd IGV
                        anusha@cn1:/raid/development/anusha/IGV/IGV/IGV> ls
                        IGV SRR203400.bowtieold.bam SRR203400.sort.bam.bam SRR203400.sort.bam.bam.bai
                        anusha@cn1:/raid/development/anusha/IGV/IGV/IGV> ls
                        IGV SRR203400.bowtieold.bam SRR203400.sort.bam.bam SRR203400.sort.bam.bam.bai
                        anusha@cn1:/raid/development/anusha/IGV/IGV/IGV> cd ..
                        anusha@cn1:/raid/development/anusha/IGV/IGV> cd ..
                        anusha@cn1:/raid/development/anusha/IGV> ls
                        IGV SRR203400.bowtieold.bam SRR203400.sort.bam.bam SRR203400.sort.bam.bam.bai
                        anusha@cn1:/raid/development/anusha/IGV> pwd
                        /raid/development/anusha/IGV
                        anusha@cn1:/raid/development/anusha/IGV> cd ..
                        anusha@cn1:/raid/development/anusha> ln -s /raid/development/anusha/SRR203400.bowtieold.bam /raid/development/anusha/SRR203400.bowtieold.bam
                        ln: failed to create symbolic link ‘/raid/development/anusha/SRR203400.bowtieold.bam’: File exists
                        anusha@cn1:/raid/development/anusha> Write failed: Broken pipe
                        Anushas-MacBook-Air:~ AnuSuri$ ssh [email protected]
                        Password:
                        Last login: Fri Sep 27 09:44:53 2013 from 172.21.164.246

                        #################################
                        Personal compute nodes : cn1-cn3
                        Scheduled compute nodes: cn4-cn19
                        #################################

                        anusha@hn:~> ls
                        50M-baits_bait.interval_list1 ANUSHA_PROJECT humanV4-baits_hg19_lite.interval_list _mappedHuman_nameSorted.bam ncbi_error_report.xml shell_script_name.sh
                        50M-targets_hg19_lite.interval_list1 bin humanV4-targets_hg19_lite.interval_list _mappedMouse_nameSorted.bam public_html
                        anusha@hn:~> cd public_html
                        anusha@hn:~/public_html> ls
                        luad_exome.bed luad_exome.copy.bed samplecov3.bedgraph samplecovhg19.chr.bedgraph
                        anusha@hn:~/public_html> cd ...
                        -bash: cd: ...: No such file or directory
                        anusha@hn:~/public_html> cd ..
                        anusha@hn:~> ln -s /raid/development/anusha/SRR203400.bowtieold.bam /raid/development/anusha/SRR203400.bowtieold.bam
                        ln: failed to create symbolic link `/raid/development/anusha/SRR203400.bowtieold.bam': File exists
                        anusha@hn:~>

                        Hi I did try to make my files HTML . I put all my files in a folder and applied command but i can't see public folder but if i try for individual files it showing file already there . Can anyone explain me what is happening and also i did not understand what is meant by click and load can anybody explain me in detail how to do IGV?

                        Comment


                        • #13
                          Originally posted by AnushaC View Post
                          anusha@cn1:/raid/development/anusha/IGV> ln -s /raid/development/anusha/IGV/SRR203400.bowtieold.bam /raid/development/anusha/IGV/SRR203400.bowtieold.bam
                          Creating a symbolic link from a file to itself is a rather odd thing to try to do. It will never work and I haven't a clue what you're trying to accomplish by even trying.

                          Originally posted by AnushaC View Post
                          anusha@cn1:/raid/development/anusha> ln -s /raid/development/anusha/SRR203400.bowtieold.bam /raid/development/anusha/SRR203400.bowtieold.bam
                          Again, this will never work and I have no clue why you're trying to do this. This has nothing to do with making an HTML page, as per your title.

                          So, onto the second computer...

                          Originally posted by AnushaC View Post
                          anusha@hn:~> ln -s /raid/development/anusha/SRR203400.bowtieold.bam /raid/development/anusha/SRR203400.bowtieold.bam
                          See above.

                          Originally posted by AnushaC View Post
                          Can anyone explain me what is happening and also i did not understand what is meant by click and load can anybody explain me in detail how to do IGV?
                          Start IGV (if you have it installed properly, just type IGV from the command line. You will then have a GUI and the instructions of "Click 'File' and then 'Load'" will make more sense. You would do well to get some help locally from someone familiar with using linux and other unix like systems as I suspect most of your issues will boil down to "basic"* computer usage.

                          *Well, as basic as using *nix-like systems can ever be considered.

                          Comment


                          • #14
                            fastq to sam want to use -F options to know only the read that are aligned to genome

                            Hi Everyone ,
                            can anyone can explain me how to use -F option for sam output to know the only the reads that are aligned to genome .



                            anusha@cn1:/raid/development/anusha> bowtie -v 3 -k 2 --sam /raid/references-and-indexes/hg19/bowtie-indexes/hg19 /raid/development/anusha/SRR203400.fastq > samtools view -S F /raid/development/anusha/SRR203400.bowtieold.flagtrnc.sam
                            Extra parameter(s) specified: "F", "/raid/development/anusha/SRR203400.bowtieold.flagtrnc.sam"
                            Command: bowtie -v 3 -k 2 --sam -S /raid/references-and-indexes/hg19/bowtie-indexes/hg19 /raid/development/anusha/SRR203400.fastq view F /raid/development/anusha/SRR203400.bowtieold.flagtrnc.sam
                            anusha@cn1:/raid/development/anusha> bowtie -v 3 -k 2 --sam /raid/references-and-indexes/hg19/bowtie-indexes/hg19 /raid/development/anusha/SRR203400.fast -F /raid/development/anusha/SRR203400.bowtieold.flagtrnc.sam
                            Command: bowtie -v 3 -k 2 --sam /raid/references-and-indexes/hg19/bowtie-indexes/hg19 /raid/development/anusha/SRR203400.fast -F /raid/development/anusha/SRR203400.bowtieold.flagtrnc.sam
                            anusha@cn1:/raid/development/anusha>

                            Comment


                            • #15
                              Checking for SNP in IGV

                              Hi Everybody ,
                              Can anyone explain me how to look for the SNP's in IGV browser


                              Thanks,
                              Anusha.Ch

                              Comment

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