Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • Predicting true SNPs from .vcf file

    The .vcf file contains so many different measurements, and a couple of different quality scores, does anyone have a good empirical idea of which values are the best guidelines to picking out real SNPs from noise and artifacts? I've done about a hundred sanger sequencing reactions on a variety of predicted SNPs at a variety of quality levels, but the picture still isn't very clear.

    For instance these SNPs looked real in the sanger:

    GT:PL:GQ 0/1:20,11,149:13 gcacacacacacacacacacacacacacacacacacacacac gcacacacacacacacacacacacacacacacacacacac 211 DP=95 DP4=17,0,10,2 MQ=48 FQ=214

    GT:PL:GQ 1/1:30,3,0:39 GAA GA 147 DP=10 DP4=0,0,1,6 MQ=48 FQ=-37.3


    These did not confirm with sanger sequencing

    GT:PL:GQ 0/1:24,0,114:27 A C 110 DP=130 DP4=14,35,2,22 MQ=45 FQ=113

    GT:PL:GQ 0/1:40,0,47:42 T G 98.3 DP=71 DP4=17,1,14,0 MQ=44 FQ=101

    Those don't look notably worse than the ones above them, so I'm not sure what I should have looked at to predict that the bottom two were false positives.

    (My a priori assumption was that these variants were all real, because I made a multi-vcf with mpileup with this samples and many sibling animals, and these variants were common to all the animals)

  • #2
    maybe the depth of the bottom two is too high. Have you try the -D options?

    Originally posted by swbarnes2 View Post
    The .vcf file contains so many different measurements, and a couple of different quality scores, does anyone have a good empirical idea of which values are the best guidelines to picking out real SNPs from noise and artifacts? I've done about a hundred sanger sequencing reactions on a variety of predicted SNPs at a variety of quality levels, but the picture still isn't very clear.

    For instance these SNPs looked real in the sanger:

    GT:PL:GQ 0/1:20,11,149:13 gcacacacacacacacacacacacacacacacacacacacac gcacacacacacacacacacacacacacacacacacacac 211 DP=95 DP4=17,0,10,2 MQ=48 FQ=214

    GT:PL:GQ 1/1:30,3,0:39 GAA GA 147 DP=10 DP4=0,0,1,6 MQ=48 FQ=-37.3


    These did not confirm with sanger sequencing

    GT:PL:GQ 0/1:24,0,114:27 A C 110 DP=130 DP4=14,35,2,22 MQ=45 FQ=113

    GT:PL:GQ 0/1:40,0,47:42 T G 98.3 DP=71 DP4=17,1,14,0 MQ=44 FQ=101

    Those don't look notably worse than the ones above them, so I'm not sure what I should have looked at to predict that the bottom two were false positives.

    (My a priori assumption was that these variants were all real, because I made a multi-vcf with mpileup with this samples and many sibling animals, and these variants were common to all the animals)

    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
    12 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, Yesterday, 06:07 PM
    0 responses
    10 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 03-22-2024, 10:03 AM
    0 responses
    52 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 03-21-2024, 07:32 AM
    0 responses
    68 views
    0 likes
    Last Post seqadmin  
    Working...
    X