Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • Library Multimerizing After PCR

    This is the most weird thing. I wonder if anyone has seen this problem:
    On Agilent DNA 1000 chip, my paired end library after PCR amplification has a little bit of high molecular weigh stuff that looks like multimers of the fragment of interest (~300bp). So I purified it to remove the high MW stuff and re-ran it on bioanalyzer but again I saw that same multimer!

    Any idea what it is? I thought it was a PCR artifact initially. If that was the case it should be gone after the purification step but it's not. Is it a real multimer?

  • #2
    It could be heteroduplexes forming during late stage PCR or during purification (esp. using chaotropic detergents s.a. guanidine salts in combination with heat). They migrate slower on gel, check section II in Kanagawa T (2003).

    Edit: This should only be prominent if you have homologous regions in your sequences. But maybe the adapter regions are long enough to bring non-identical DNA strands together.
    Last edited by sulfobus; 04-20-2010, 03:50 AM.

    Comment

    Latest Articles

    Collapse

    • seqadmin
      Essential Discoveries and Tools in Epitranscriptomics
      by seqadmin




      The field of epigenetics has traditionally concentrated more on DNA and how changes like methylation and phosphorylation of histones impact gene expression and regulation. However, our increased understanding of RNA modifications and their importance in cellular processes has led to a rise in epitranscriptomics research. “Epitranscriptomics brings together the concepts of epigenetics and gene expression,” explained Adrien Leger, PhD, Principal Research Scientist...
      04-22-2024, 07:01 AM
    • seqadmin
      Current Approaches to Protein Sequencing
      by seqadmin


      Proteins are often described as the workhorses of the cell, and identifying their sequences is key to understanding their role in biological processes and disease. Currently, the most common technique used to determine protein sequences is mass spectrometry. While still a valuable tool, mass spectrometry faces several limitations and requires a highly experienced scientist familiar with the equipment to operate it. Additionally, other proteomic methods, like affinity assays, are constrained...
      04-04-2024, 04:25 PM

    ad_right_rmr

    Collapse

    News

    Collapse

    Topics Statistics Last Post
    Started by seqadmin, Yesterday, 08:47 AM
    0 responses
    12 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 04-11-2024, 12:08 PM
    0 responses
    60 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 04-10-2024, 10:19 PM
    0 responses
    59 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 04-10-2024, 09:21 AM
    0 responses
    54 views
    0 likes
    Last Post seqadmin  
    Working...
    X