Announcement

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

  • Increasing MiSeq v3 cycle number for >600bp reads

    Hello people!

    We have a sequencing application where we could really need, say, 50–100 more cycles from a 600-cycle MiSeq v3 kit.

    Illumina states that the cartridge contains reagents for 625 cycles [1] (to account for dual indices we don't need, so 10–15 more cycles there already).
    Then again, ECO got about 370 cycles from a 300 cycle kit [2] which is a fair bit more than the 325 stated by Illumina. Other people also mentioned that they pushed above the 25 reserve index cycles [3].

    Lastly, this post explains how MadsAlbertsen got 600 cycles from a 400 cycle kit by filling up some wells with reagents from leftover cartridges.

    So, my question: Does anybody has experience with increasing the cycle number on the 600-cycle v3 kits and can tell me what safely works? Either by experimenting how many possible cycles are in one cartrige or by filling up from an old cartridge.

  • #2
    People have been doing similar things on the HiSeq too - but the issue here isn't whether it's technically feasible to hack the kits, it's whether your application will be able to cope with the awful sequencing quality of anything over 300bp in length. If you routinely use the 600 cycle kits, you'll know how the R2 drop off is - so imagine what this is going to be like after 700 cycles...

    But see also: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3673215/

    Comment


    • #3
      Thanks, what they did in the paper looks really cool. And yes, I am aware of the decreasing quality, but a small amount of data (after filtering) helps us more than none, in this case...

      Comment


      • #4
        Have you considered using nanopore sequencing if you need long(er) reads? No hacking of sequencer needed.

        Comment


        • #5
          Not for this, as it's an antibody library and we need a single paired read with low error rate to cover one fragment. Hacking it will probably be

          Comment


          • #6
            If you do not need high number of reads, PacBio CCS will be a good option as it will cover the whole fragment length with very high accuracy.

            Comment

            Latest Articles

            Collapse

            • seqadmin
              Advanced Tools Transforming the Field of Cytogenomics
              by seqadmin


              At the intersection of cytogenetics and genomics lies the exciting field of cytogenomics. It focuses on studying chromosomes at a molecular scale, involving techniques that analyze either the whole genome or particular DNA sequences to examine variations in structure and behavior at the chromosomal or subchromosomal level. By integrating cytogenetic techniques with genomic analysis, researchers can effectively investigate chromosomal abnormalities related to diseases, particularly...
              09-26-2023, 06:26 AM
            • seqadmin
              How RNA-Seq is Transforming Cancer Studies
              by seqadmin



              Cancer research has been transformed through numerous molecular techniques, with RNA sequencing (RNA-seq) playing a crucial role in understanding the complexity of the disease. Maša Ivin, Ph.D., Scientific Writer at Lexogen, and Yvonne Goepel Ph.D., Product Manager at Lexogen, remarked that “The high-throughput nature of RNA-seq allows for rapid profiling and deep exploration of the transcriptome.” They emphasized its indispensable role in cancer research, aiding in biomarker...
              09-07-2023, 11:15 PM

            ad_right_rmr

            Collapse

            News

            Collapse

            Topics Statistics Last Post
            Started by seqadmin, Yesterday, 09:38 AM
            0 responses
            9 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 09-27-2023, 06:57 AM
            0 responses
            11 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 09-26-2023, 07:53 AM
            1 response
            23 views
            0 likes
            Last Post seed_phrase_metal_storage  
            Started by seqadmin, 09-25-2023, 07:42 AM
            0 responses
            17 views
            0 likes
            Last Post seqadmin  
            Working...
            X