Seqanswers Leaderboard Ad

Collapse

Announcement

Collapse
No announcement yet.
X
 
  • Filter
  • Time
  • Show
Clear All
new posts

  • Anyone has experience with Bowtie2?

    Hi All,
    my name is Lucia, and I've been lurking around for a while but this is my first post.
    I have a pretty big ChIP-seq experiment to analyze, and initially did the alignments with Bowtie. The stats were not that great, and since I have 100bp reads, I decided to redo the alignments with Bowtie2
    I do get a lot more mapped reads, but the quality scores seem a lot lower
    The programs compute the scores differently, so I am not sure if the scores are really "lower".
    Does Bowtie2 increase mapping at the expense of quality?
    Or is the scoring scheme so completely different that the quality scores are not comparable at all?
    thanks

  • #2
    Sorry can't help you with your specific problem.

    Though in my experience, bowtie works much better than bowtie2. Apart from some bugs, many options are missing (strata/unique alignments). I shared my opinion with authors as well.
    Must mention that I have 50bp reads and my final goal (ChIP-Seq) may be different from you.

    Comment


    • #3
      Maybe try bwa

      Comment


      • #4
        Depends on what score you are talking about. The MAPQ score from bowtie2 has to do with the uniquness of the alignment and not necessarily the quality of the alignment. You want to look at the AS attribute in the extended attributes of the SAM files. Also read up on this section of the bowtie2 manual:



        By default bowtie2 is allowing no mismatches in the seed (22 bases for default end-to-end alignment) and a minimum alignment score of -60.6 (with a max of 0) for 100bp reads. Default penalties are something like -6 for a mismatch at a high quality base an -11 for a length-2 gap. Gaps in reads and reference are scored the same. Specifically you get a -5 for a gap open and -3 for each base. So a length-1 gap is -8, length-2 is -11, etc. So with a min score of about -60 you could have as many as 6 mismatches past the seed or all kinds of combinations of gaps and mismatches. You can control all of those parameters. The min score setting is defined as -0.6 + -0.6*L where L is you read length. If you want to set that value you do so with --min-score L,-0.6,-0.6. So you can figure out what you want your min score to be, like say -40, and work backward from that to figure out what values to put in the setting.
        /* Shawn Driscoll, Gene Expression Laboratory, Pfaff
        Salk Institute for Biological Studies, La Jolla, CA, USA */

        Comment


        • #5
          Mapping quality is valuable for variant calling. If the quality score does not correspond to accuracy of alignment, this is a big problem I think.

          Comment


          • #6
            Originally posted by adaptivegenome View Post
            Mapping quality is valuable for variant calling. If the quality score does not correspond to accuracy of alignment, this is a big problem I think.
            You're right about that. That's probably why it's better to use BWA for those alignments. I don't know of any advantage other than speed for using bowtie2 over BWA. I haven't done any hardcore comparisons between the two. I do like BWA's alignments more though...not sure why.
            /* Shawn Driscoll, Gene Expression Laboratory, Pfaff
            Salk Institute for Biological Studies, La Jolla, CA, USA */

            Comment

            Latest Articles

            Collapse

            • seqadmin
              Current Approaches to Protein Sequencing
              by seqadmin


              Proteins are often described as the workhorses of the cell, and identifying their sequences is key to understanding their role in biological processes and disease. Currently, the most common technique used to determine protein sequences is mass spectrometry. While still a valuable tool, mass spectrometry faces several limitations and requires a highly experienced scientist familiar with the equipment to operate it. Additionally, other proteomic methods, like affinity assays, are constrained...
              04-04-2024, 04:25 PM
            • seqadmin
              Strategies for Sequencing Challenging Samples
              by seqadmin


              Despite advancements in sequencing platforms and related sample preparation technologies, certain sample types continue to present significant challenges that can compromise sequencing results. Pedro Echave, Senior Manager of the Global Business Segment at Revvity, explained that the success of a sequencing experiment ultimately depends on the amount and integrity of the nucleic acid template (RNA or DNA) obtained from a sample. “The better the quality of the nucleic acid isolated...
              03-22-2024, 06:39 AM

            ad_right_rmr

            Collapse

            News

            Collapse

            Topics Statistics Last Post
            Started by seqadmin, 04-11-2024, 12:08 PM
            0 responses
            17 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 04-10-2024, 10:19 PM
            0 responses
            21 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 04-10-2024, 09:21 AM
            0 responses
            16 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 04-04-2024, 09:00 AM
            0 responses
            46 views
            0 likes
            Last Post seqadmin  
            Working...
            X