Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • bwa mem - low properly paired percentage

    After aligning paired-end 100bp reads to a reference genome, I am getting very low properly paired percentage:

    369208441 0 total (QC-passed reads + QC-failed reads)
    8985531 0 secondary
    289733341 0 mapped
    78.47% N/A mapped %
    360222910 0 paired in sequencing
    180111455 0 read1
    180111455 0 read2
    1393338 0 properly paired
    0.39% N/A properly paired %
    280747810 0 with itself and mate mapped
    0 0 singletons
    0.00% N/A singletons %
    39590468 0 with mate mapped to a different chr
    0 0 with mate mapped to a different chr (mapQ>=5)

    I followed GATK best practices to align paired-end short-read data to a reference genome. I downloaded the short-read data from NCBI SRA into fastq files using SRA toolkit's fastq-dump, converted the fastq files into unmapped bam using Picard FastqToSam, and marked adapters using Picard MarkIlluminaAdapters. I then piped Picard SamToFastq, bwa mem, and Picard MergeBamAlignment. To get stats on the alignment, I used samtools flagstat. For several of my samples, the alignment went great (90% mapped, 80% properly paired). However, for a couple of my samples, the properly paired percentage was well below 1%. I'm wondering how I could have a normal amount of reads mapping (~78%) but have only .39% of those reads properly paired.

    I have double-checked that my fastq files from fastq-dump have identical read counts, and that they are properly interleaved after Picard FastqToSam.

  • #2
    Cross-posted at biostars: https://www.biostars.org/p/464101/

    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, Today, 06:37 PM
    0 responses
    6 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, Today, 06:07 PM
    0 responses
    6 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