Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • NGS result that can't be verified with RT-PCR

    I received 5 plant leaf samples for NGS. Two of the 5 were determined to be positive for virus via RT-PCR. However when I performed NGS, all 5 samples were determined to be positive. In the 3 samples that were supposed to be negative, I acquired complete genomes when assembled using CLC Genomics Workbench.

    I can't seem to find any source of contamination. The 3 supposed negative samples are of a different strain than the other two. However, it is a strain that should still be picked up by my RT-PCR primers.

    I hadn't used these indices in a while as I know that I have had problems with false positives in the past when using the same index sequences in consecutive runs. This is why I always rotate my index sequences for each NGS run. The last several times these index sequences were used, they did not detect that target. So, I'm thinking that it's not that type of contamination.

    I've ruled out that it's pipette contamination because I'm using the same pipettes on library prep as I do with RT-PCR.

    The run was a near perfect 1452Kmm-2. So, overclustering leading to strange base calls can also be ruled out.

    Does anyone have any ideas what could be going on here?

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
10 views
0 likes
Last Post seqadmin  
Started by seqadmin, Yesterday, 06:07 PM
0 responses
9 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
67 views
0 likes
Last Post seqadmin  
Working...
X