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  • Hi, Ben Langmead
    Recently I used bowtie to align some solexa reads in 100bp length. But the matched ratio is low. I want to know what's your recommendation for the option parameters for the long reads such as 100bp?

    Thank you

    Comment


    • Hi Ben, I am a very new beginner with bowtie. I have a question about it, hope you can help me out. When I run bowtie with 454 long reads(the read length is about 200bp), the result is very weird that nearly 98% of reads are mismatching. (I tried human, yeast and e_coli). Could you tell me that whether I can use bowtie to align the long read or not....Thx

      Comment


      • Originally posted by cp229 View Post
        Hi Ben, I am a very new beginner with bowtie. I have a question about it, hope you can help me out. When I run bowtie with 454 long reads(the read length is about 200bp), the result is very weird that nearly 98% of reads are mismatching. (I tried human, yeast and e_coli). Could you tell me that whether I can use bowtie to align the long read or not....Thx
        The 454 reads may contain indel errors while sequencing. Also, longer reads may cover more indel variants in the reference genome. I think the indels could be the main reason why bowtie reported a low mappable ratio.
        Xi Wang

        Comment


        • Is it possible to get mapping quality and read quality (a single value per read, e.g. calculated out of base quality) for Bowtie?

          Comment


          • Originally posted by Ben Langmead View Post
            I think the problem is that you're assuming -n 0 -l 16 is going to allow any alignment with no mismatches in the first 16 colors, but the -e limit also applies. -e also needs to be set high enough so that the sum of the quality values of all the mismatched positions in your example are still <= the -e limit. If no quality values are supplied, qualities all = 30.

            Hope that helps. You may also want to consider using trimming (e.g. -3).

            Thanks,
            Ben
            Thank you Ben! and sorry for late response...
            Now it works as I want
            Best!

            tomek

            Comment


            • I think I`ve got that now!!Thank you very much Xi Wang.

              Comment


              • Hi all

                I got almost the same problem with bowtie. I tried to align paired end read in fastaQ format of 84 bp
                >bowtie -q -t --solexa1.3-quals -p 2 --sam xx_index.txt -1 xx.txt -2 xx.txt -m 1 > xx.sam

                but I got the message
                "Reads file contained a pattern with more than 1024 sequence characters.
                Please truncate reads and quality values and and re-run Bowtie
                terminate called after throwing an instance of 'int'
                Aborted"

                but if I'm using one file of the paired end reads as single end for alignment it works

                There is no uncalled bases in my reads
                Any idea about this issue ?

                Thank you

                AC

                Comment


                • Originally posted by yeb3czg View Post
                  Hi all

                  I got almost the same problem with bowtie. I tried to align paired end read in fastaQ format of 84 bp
                  >bowtie -q -t --solexa1.3-quals -p 2 --sam xx_index.txt -1 xx.txt -2 xx.txt -m 1 > xx.sam

                  but I got the message
                  "Reads file contained a pattern with more than 1024 sequence characters.
                  Please truncate reads and quality values and and re-run Bowtie
                  terminate called after throwing an instance of 'int'
                  Aborted"

                  but if I'm using one file of the paired end reads as single end for alignment it works

                  There is no uncalled bases in my reads
                  Any idea about this issue ?

                  Thank you

                  AC
                  Have you try to put the option -m ahead, like this?

                  Code:
                  bowtie  -m 1 -q -t --solexa1.3-quals -p 2 --sam xx_index.txt -1 xx.txt -2 xx.txt > xx.sam
                  Xi Wang

                  Comment


                  • Xi
                    I tried it didn't change anything but I found what was the problem in one of my reads file the format of one lane was not correct
                    now it 's working
                    thanks

                    Comment


                    • bowtie and solid

                      Hi,
                      I have another question about solid data...:
                      I have two different reads:
                      T32221113212031121021022123023302010
                      and
                      T22221113212031121021022123023302010
                      In color space difference is only in one number - so in the base space those sequences are completely different, however with bowtie they are mapped to the same seq:

                      bowtie -a -n 0 -C ../indeks/miRNA-mature_cs -3 8 -c T32221113212031121021022123023302010
                      0 + mmT-miR-1944 1 TCTGTGCTGAATGTCAAGTTCTGAT qqqqqqqqqqqqqqqqqqqqqqqqq 0


                      bowtie -a -n 0 -C ../indeks/miRNA-mature_cs -3 8 -c T22221113212031121021022123023302010
                      0 + mmT-miR-1944 1 TCTGTGCTGAATGTCAAGTTCTGAT qqqqqqqqqqqqqqqqqqqqqqqqq 0

                      Why?
                      Thanks for any suggestions!
                      Best!

                      tomek

                      Comment


                      • Originally posted by didymos View Post
                        Hi,
                        I have another question about solid data...:
                        I have two different reads:
                        T32221113212031121021022123023302010
                        and
                        T22221113212031121021022123023302010
                        In color space difference is only in one number - so in the base space those sequences are completely different, however with bowtie they are mapped to the same seq:

                        bowtie -a -n 0 -C ../indeks/miRNA-mature_cs -3 8 -c T32221113212031121021022123023302010
                        0 + mmT-miR-1944 1 TCTGTGCTGAATGTCAAGTTCTGAT qqqqqqqqqqqqqqqqqqqqqqqqq 0


                        bowtie -a -n 0 -C ../indeks/miRNA-mature_cs -3 8 -c T22221113212031121021022123023302010
                        0 + mmT-miR-1944 1 TCTGTGCTGAATGTCAAGTTCTGAT qqqqqqqqqqqqqqqqqqqqqqqqq 0

                        Why?
                        Thanks for any suggestions!
                        Best!

                        tomek
                        For one of the two sequences, the first color was probably identified as a color error (sequencing error) and corrected appropriately. Color errors (sequencing error) and base differences (SNPs) manifest differently. See the attached PDF for a brief explanation between the differences.
                        Attached Files

                        Comment


                        • Originally posted by nilshomer View Post
                          For one of the two sequences, the first color was probably identified as a color error (sequencing error) and corrected appropriately. Color errors (sequencing error) and base differences (SNPs) manifest differently. See the attached PDF for a brief explanation between the differences.
                          Thank you!
                          very clear presentation. However in my case things are little bit different.
                          I am mapping not to the whole genome but only to the miRNA sequences (previously indexed with bowtie-build). If I understand correctly color is identified as color error when after correction sequence can be easily mapped, but in my case this "error" sequence can be just not a miRNA sequence - other RNA type from RNA-seq experiment.
                          Question is how many color errors are allowed to be correct in bowtie - how I can set it or how can I switch it off if possible?
                          Thanks!

                          tomek

                          Comment


                          • Missed alignment?

                            I noticed that sometimes when I use Bowtie, if the only mismatch is at the 5' end, then no alignment will be produced. Could this be a bug from index building? Here is an example below. With only a single mismatch at the very 5' end, I expect to see the read aligned with -v 2. However, that is not the case. Could someone tell me why and how this can be corrected? Many thanks in advance.

                            ../bowtie-0.12.5/bowtie -t refseq_hs -c CGACTCTTAGCGGTGGATCACTCGG -v 2
                            # reads processed: 1
                            # reads with at least one reported alignment: 0 (0.00%)
                            # reads that failed to align: 1 (100.00%)
                            No alignments


                            ../bowtie-0.12.5/bowtie -t refseq_hs -c GACTCTTAGCGGTGGATCACTCGG -v 2
                            0 + gi|142372596|ref|NR_003285.2| 0 GACTCTTAGCGGTGGATCACTCGG IIIIIIIIIIIIIIIIIIIIIIII 0
                            # reads processed: 1
                            # reads with at least one reported alignment: 1 (100.00%)
                            # reads that failed to align: 0 (0.00%)
                            Reported 1 alignments to 1 output stream(s)

                            Comment


                            • Originally posted by laghs View Post
                              I noticed that sometimes when I use Bowtie, if the only mismatch is at the 5' end, then no alignment will be produced. Could this be a bug from index building? Here is an example below. With only a single mismatch at the very 5' end, I expect to see the read aligned with -v 2. However, that is not the case. Could someone tell me why and how this can be corrected? Many thanks in advance.

                              ../bowtie-0.12.5/bowtie -t refseq_hs -c CGACTCTTAGCGGTGGATCACTCGG -v 2
                              # reads processed: 1
                              # reads with at least one reported alignment: 0 (0.00%)
                              # reads that failed to align: 1 (100.00%)
                              No alignments


                              ../bowtie-0.12.5/bowtie -t refseq_hs -c GACTCTTAGCGGTGGATCACTCGG -v 2
                              0 + gi|142372596|ref|NR_003285.2| 0 GACTCTTAGCGGTGGATCACTCGG IIIIIIIIIIIIIIIIIIIIIIII 0
                              # reads processed: 1
                              # reads with at least one reported alignment: 1 (100.00%)
                              # reads that failed to align: 0 (0.00%)
                              Reported 1 alignments to 1 output stream(s)
                              I think your case was due to Bowtie cann't deal with indel, but not because of 5' mismatches. Note that the aligned position for read GACTCTTAGCGGTGGATCACTCGG (your 2nd read) is 0 (0-based), so for your 1st read: CGACTCTTAGCGGTGGATCACTCGG, it was not able to map to the reference sequences.
                              Xi Wang

                              Comment


                              • Originally posted by Xi Wang View Post
                                I think your case was due to Bowtie cann't deal with indel, but not because of 5' mismatches. Note that the aligned position for read GACTCTTAGCGGTGGATCACTCGG (your 2nd read) is 0 (0-based), so for your 1st read: CGACTCTTAGCGGTGGATCACTCGG, it was not able to map to the reference sequences.
                                I see. Is there a way to get around this problem (other than using another alignment program)? Thanks.

                                Comment

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