Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • Nextera exome kit

    Hi:
    Recently we tested the nextera exome enrichment libe kit. In our analysis pipeline we have a metric that calculates exonic reads, intronic reads, and intergenic reads. Our nextera libes all had about 50% match to the exons. That seemed low, I went back and looked at the same metric from the original illumina kits, those also had only about 50% exonic reads. Does this seem low? It throws a real wrench into coverage calculations

  • #2
    What is the on-target specification of Nextera? I don't use Nextera, I use SureSelect, which will generally give ~65% 'on target' reads on variable quality samples, but the Agilent literature claims 70%. For custom capture regions on SureSelect, 50% on target is all you will get, so it depends. Also are you feeding a list of all exons to your coverage metric calculator? Or just the exons in the capture kit?

    Comment


    • #3
      Thanks for your comments. That is an excellent point, I'm sure we are feeding all known exons so that could be a big part of it. I'll know a lot more when we send the sureselect material info through the same pipeline, but it does sound as though those numbers are to be expected. Just looking around on the browser it is clear that are abundant areas of off target seqeuncing.

      Comment


      • #4
        We've outlined each currently available capture technology in a comparison table here:
        Whole exome sequencing services guide compares exome capture kits, demonstrates how to calculate the reads required to meet coverage requirements and makes best practice recommendations.


        In our guide for determining the number of sequencing reads one needs for a targeted capture expt, we estimate on-target rates to be between 65-75%. I'd be interested in hearing other people's experiences and their rates.

        - Genohub

        Comment


        • #5
          In my lab we are using the Nextera Rapid Capture Exome kit and I also get only slightly above 50% of all reads mapping on the target region. Unfortunately, this number has been low and stable over multiple runs. In my experience, the on-target or coverage estimations given by the vendors are always to high and cannot be reproduced.

          Comment

          Latest Articles

          Collapse

          • 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
          • seqadmin
            Techniques and Challenges in Conservation Genomics
            by seqadmin



            The field of conservation genomics centers on applying genomics technologies in support of conservation efforts and the preservation of biodiversity. This article features interviews with two researchers who showcase their innovative work and highlight the current state and future of conservation genomics.

            Avian Conservation
            Matthew DeSaix, a recent doctoral graduate from Kristen Ruegg’s lab at The University of Colorado, shared that most of his research...
            03-08-2024, 10:41 AM

          ad_right_rmr

          Collapse

          News

          Collapse

          Topics Statistics Last Post
          Started by seqadmin, Yesterday, 06:37 PM
          0 responses
          7 views
          0 likes
          Last Post seqadmin  
          Started by seqadmin, Yesterday, 06:07 PM
          0 responses
          7 views
          0 likes
          Last Post seqadmin  
          Started by seqadmin, 03-22-2024, 10:03 AM
          0 responses
          49 views
          0 likes
          Last Post seqadmin  
          Started by seqadmin, 03-21-2024, 07:32 AM
          0 responses
          66 views
          0 likes
          Last Post seqadmin  
          Working...
          X