Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • IDT xGen kit on RNAseq libraries?

    I'm looking into using the xGen lockdown kit to select for a panel of specific genes from our RNA seq samples. These are low input samples, so my normal workflow is doing about 13 cycles of preamp, then pool and SPRI clean. Then I take that into Nextera, which is then amplified again with the Nextera handles for 12 cycles. This usually produced me with sufficient DNA for a NextSeq run (1-4nM, maybe about 10-20uL total).

    I'm looking into doing a gene enrichment with the xGEN kit after the preamp. but the recommended input of the kit seems really high. It asks for 500ng of material, while my preamp material is usually around 1-2ng/uL (15uL total). Does anybody have any idea whether the kit would still work with much lower input? Or is there a different kit that would be more suited for this purpose? (the libraries are already barcoded. so no additional barcoding is necessary.)

  • #2
    I've tried the kit with lower input from stranded RNA library preps and it seems to be fine at low coverage (~40X). i'd probably try to stay above 150-200ng for the hybs if you're trying to cut down on input. If duplication rates become an issue you might want to increase input, but it will depend on how deep you want to sequence and on the complexity of your library, pre-cap.

    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 on Modified Bases...
      Yesterday, 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, 04-11-2024, 12:08 PM
    0 responses
    55 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 04-10-2024, 10:19 PM
    0 responses
    51 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 04-10-2024, 09:21 AM
    0 responses
    45 views
    0 likes
    Last Post seqadmin  
    Started by seqadmin, 04-04-2024, 09:00 AM
    0 responses
    55 views
    0 likes
    Last Post seqadmin  
    Working...
    X