Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • Reduce PhiX

    Dear all,

    I was wondering if there was any tool to help me reduce the PhiX% used in our NextSeq 500 runs? We run similar samples continuously, with "low" diversity as we enrich specific regions by hybrid capture. Instead of empirical & successive testings to reduce the PhiX%, can I guestimate the "ideal" PhiX% to add before reaching the slippery slope?
    Thank you for your help.
    Gaetan

  • #2
    Hi Gaetan,

    For sequencing on the MiSeq at least I know there is a recommendation to use a 5% PhiX spike-in for low diversity libraries (dependent on which version of the MiSeq Control Software you are using, you'll have to look that up.)

    When I sequenced on the NextSeq I also used a hybridisation/capture approach however this was exome capture and the diversity was fine so I used a 1% PhiX spike-in for QC purposes.

    For your purposes, a 5% minimum PhiX spike-in would probably do the trick, but I may stand corrected if someone else chimes in.

    Cheers!

    Comment


    • #3
      Hello,

      Thank you for your quick answer!

      Unfortunately, I cannot test right down to 5% as my samples are way to precious to risk a failed run because of low diversity.

      So there no tools/tests to gauge the diversity based on my collected data that could help me to reduce the "wasted" output in sequencing "too much" PhiX?

      I could safely slowly reduce the PhiX% each run until I see a deterioration in quality metrics but this empirical testing sounds like a waste of time if I am far off...

      Thank you in advance!

      Cheers

      Comment


      • #4
        Hi Gaetan,

        Yeah it's always a bother when working with precious samples and NGS! I took a quick look and found this from August 2018:



        The NextSeq guidelines for PhiX spike-ins for low diversity libraries is 10% - 50%. I would be relatively confident that a 10% spike-in would be a good starting point. If you are still hesitant, increase that to 15% - 20%.


        For your other question - unfortunately I am not aware of any tool to deduce diversity I'm afraid. Sorry!

        Cheers.

        Comment


        • #5
          Thank you again for your even quicker response!

          I got those numbers too, but I guess it depends a lot on your target panel size, dups, or depth required for your analysis...

          I will keep digging and if I found something, I'll post it!

          Thanks again!

          Comment

          Latest Articles

          Collapse

          • 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
          • seqadmin
            The Impact of AI in Genomic Medicine
            by seqadmin



            Artificial intelligence (AI) has evolved from a futuristic vision to a mainstream technology, highlighted by the introduction of tools like OpenAI's ChatGPT and Google's Gemini. In recent years, AI has become increasingly integrated into the field of genomics. This integration has enabled new scientific discoveries while simultaneously raising important ethical questions1. Interviews with two researchers at the center of this intersection provide insightful perspectives into...
            02-26-2024, 02:07 PM

          ad_right_rmr

          Collapse

          News

          Collapse

          Topics Statistics Last Post
          Started by seqadmin, 03-14-2024, 06:13 AM
          0 responses
          34 views
          0 likes
          Last Post seqadmin  
          Started by seqadmin, 03-08-2024, 08:03 AM
          0 responses
          72 views
          0 likes
          Last Post seqadmin  
          Started by seqadmin, 03-07-2024, 08:13 AM
          0 responses
          82 views
          0 likes
          Last Post seqadmin  
          Started by seqadmin, 03-06-2024, 09:51 AM
          0 responses
          68 views
          0 likes
          Last Post seqadmin  
          Working...
          X