Seqanswers Leaderboard Ad

Collapse

Announcement

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

  • Question about solexa sequencing

    Hi all,

    I am a new here, hope everyone can have a nice day!

    Here is my question, before sequencing, we have to make some libraries in different insert size. Is that the short insert size (200 ~500 bp) are using single-end sequencing for the alignment of contigs, and the long insert size (over 2kb) are using paie-end sequencing for joining the contigs into scaffolds?

    Thanks in advance

    Man

  • #2
    If you're using the Illumina platform for sequencing then your choice of insert size is limited by the technique used to generate the clusters on the flowcell. Generally your inserts have to be <800bp to produce usable clusters so you don't really have the option to use longer inserts.

    Comment


    • #3
      So, if I want to sequence a species, is it neccessary to senquence the single-end or what I can do is just the pair-end sequencing then align the reads?

      Thanks

      Comment


      • #4
        Originally posted by simonandrews View Post
        If you're using the Illumina platform for sequencing then your choice of insert size is limited by the technique used to generate the clusters on the flowcell. Generally your inserts have to be <800bp to produce usable clusters so you don't really have the option to use longer inserts.
        This is true for the Illumina paired-end protocol, in which you sequence both ends of a contiguous stretch to DNA. But Illumina also has the mate-pair protocol which allows you to get paired reads separated by 2-5kbp.

        Comment


        • #5
          Originally posted by kmcarr View Post
          This is true for the Illumina paired-end protocol, in which you sequence both ends of a contiguous stretch to DNA. But Illumina also has the mate-pair protocol which allows you to get paired reads separated by 2-5kbp.
          Anyone cares to talk about its experience with that protocol? How stable is it?
          -drd

          Comment


          • #6
            Originally posted by drio View Post
            Originally posted by kmcarr View Post
            This is true for the Illumina paired-end protocol, in which you sequence both ends of a contiguous stretch to DNA. But Illumina also has the mate-pair protocol which allows you to get paired reads separated by 2-5kbp.
            Anyone cares to talk about its experience with that protocol? How stable is it?
            We have done it once and it mostly worked as expected. That is to say the vast majority of reads were properly paired and spaced ~2.8-3.0 kbp apart. However we observed and extreme bias in the nucleotide composition at the start of the reads. I posted about this in another thread (see here) but unfortunately got no hits there. I to am very interested in hearing what other's experiences have been with the mate-pair protocol.

            Comment


            • #7
              Originally posted by kmcarr View Post
              We have done it once and it mostly worked as expected. That is to say the vast majority of reads were properly paired and spaced ~2.8-3.0 kbp apart. However we observed and extreme bias in the nucleotide composition at the start of the reads. I posted about this in another thread (see here) but unfortunately got no hits there. I to am very interested in hearing what other's experiences have been with the mate-pair protocol.
              I'll get back to you on that.. I have to double check.
              Ping me if I don't reply in the next couple of days.
              I'll reply to your other post.
              -drd

              Comment

              Latest Articles

              Collapse

              • 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
              • 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

              ad_right_rmr

              Collapse

              News

              Collapse

              Topics Statistics Last Post
              Started by seqadmin, 04-11-2024, 12:08 PM
              0 responses
              22 views
              0 likes
              Last Post seqadmin  
              Started by seqadmin, 04-10-2024, 10:19 PM
              0 responses
              24 views
              0 likes
              Last Post seqadmin  
              Started by seqadmin, 04-10-2024, 09:21 AM
              0 responses
              19 views
              0 likes
              Last Post seqadmin  
              Started by seqadmin, 04-04-2024, 09:00 AM
              0 responses
              50 views
              0 likes
              Last Post seqadmin  
              Working...
              X