Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • sequencing QC and metagenomics

    Hi,

    I just started my first metagenomics shot-gun sequencing analysis. I hope I can get some advice here.

    1. QC process: I am looking at removing duplicate sequences and trimming low quality bases, according to this HMP's SOP. Is this adequate? What is considered a good QC practice?

    2. Data analysis: I am first trying MetaPhlAn by Huttenhower's group. Is there any 'standard' procedure/workflow that this community recommends?

    Thanks for your help,
    John

  • #2
    Originally posted by JQL View Post
    1. QC process: I am looking at removing duplicate sequences and trimming low quality bases, according to this HMP's SOP. Is this adequate? What is considered a good QC practice?
    You may want to remove human/host DNA as well. kneaddata from Curtis Huttonhower's group does this, but uses bowtie2's --un-conc output to filter reads, which misses repetitive DNA that doesn't align concordantly. We've taken to manually filtering any read pair that aligns in any fashion to the human reference.

    Originally posted by JQL View Post
    2. Data analysis: I am first trying MetaPhlAn by Huttenhower's group. Is there any 'standard' procedure/workflow that this community recommends?
    There are also k-mer based approaches (e.g. kraken and CLARK) that were recently shown to outperform MetaPhlAn: see here. Generally speaking, I don't think shotgun is as well figured out as 16S, so there is still some exploring to do. You may also be interested in PanPhlAn, ConStrains, etc. that do strain-level profiling.

    We've just gotten into shotgun too, so happy to share any advice.

    Comment


    • #3
      Thank you very much for your reply. At this point, any thing will help me get started.

      have a nice weekend,
      John

      Originally posted by fanli View Post
      You may want to remove human/host DNA as well. kneaddata from Curtis Huttonhower's group does this, but uses bowtie2's --un-conc output to filter reads, which misses repetitive DNA that doesn't align concordantly. We've taken to manually filtering any read pair that aligns in any fashion to the human reference.



      There are also k-mer based approaches (e.g. kraken and CLARK) that were recently shown to outperform MetaPhlAn: see here. Generally speaking, I don't think shotgun is as well figured out as 16S, so there is still some exploring to do. You may also be interested in PanPhlAn, ConStrains, etc. that do strain-level profiling.

      We've just gotten into shotgun too, so happy to share any advice.

      Comment


      • #4
        Other things you'll need to do:
        - assembly, best cross-assembly if you have multiple samples
        - genomic binning (e.g. maxBin)
        - gene prediction (prodigal has a setting for meta, but only bacteria; ncRNAs can be predicted with rnammer, trnascan-SE and rfam)
        - function prediction (via KO, InterproScan, PRIAM, dbCAN, etc)

        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
        8 views
        0 likes
        Last Post seqadmin  
        Started by seqadmin, Yesterday, 06:07 PM
        0 responses
        8 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