Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • Contamination with bacterial rRNA

    We have done Ribo-Zero-Human rRNA depletion of total-RNA from human clinical samples followed by ScriptSeq library prep and HiSeq sequencing. But it turned out that app 50 % of the reads were of bacterial rRNA origin.
    Anyone observed something similar?
    /Jakob

  • #2
    Ribo Zero combination H/MR and bacteria

    Hello Jakob,

    The "fix" actually is simple, and it would involve using the RZ Kits in tandem. After the total RNA depletion, you can use RZ Human with the RZ bacteria (three kinds available - Gram Positive, Gram negative and "meta-Bacterial" for both gram positive and negative) in a single step (mixing the rRNA removal solutions in a 50/50 mix) or running the depletions sequentially.

    For the "combined" reaction, after total RNA purification simply use a mix of the RZ Human rRNA removal soln with the bacterial rRNA removal soln of choice. Use 5 µl of the human and 5 µl of the bacterial (total volume of 10 µl for 5 µg input). Them simply continue after the incubation at 50C, and follow the bead-binding and filtration as per normal.

    Only caveat here is to not use more than 2.5 µg of total "mixed" sample RNA (due to the overall reduction in the rRNA removal solution from what the standard volume is used in a "normal" depletion.

    If you need further clarification, do contact me.

    olaf

    Epicentre tech support

    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
    11 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
    51 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