Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • Different genomes, different libraries

    Hi everybody,

    do you have a some experience to sequence different genomes prepare with different kit in one run on MiSeq?

    We are planning to sequence about 40 BAC clones prepared by TruSeq PCR-Free, each BAC has 110 kbp, the one genome with size 120 Mbp and one genome with 3 Gbp, both prepared with Nextera DNA kit.

    I am not sure how to pooled this library (molarity) to have similar coverage for each sample. Two bigger genomes should have lower coverage.

    Any Idea?
    Thanks

  • #2
    It's ok to pool different libraries, although you'll have to demultiplex separately if the libraries have different indexing. If you want to get more coverage for your libraries of larger size, then I'd advise pooling them in adequate proportions.

    What concerns me more is sequencing 3Gbp genome on Miseq, the instrument is not purposed to sequence genomes of that size.

    40 BAC clones of 110 kbp is ~4,5Mbp in total + 120 Mbp Genome is 124,5 MBp. That should be fine to sequence on a miseq, but you might have troubles geting enough coverage for your BAC clones since most of the flowcell will probably be occupied by the 120 MBp Genome. I'd advise pooling 40 BAC clones in equalmolar, then pooling the resulting library with 120 Mbp genome 1:25 (1 part 40 clones pooled library, 25 parts Mbp genome).

    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...
      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
    52 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