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  • Leading and Lagging strand dephasing in sequencing.

    Hi,
    I'm studying the sequencing method as used by different sequenceing technology but i didn't understand the dephasing of the leading and lagging strands

  • #2
    Dephasing in sequencing usually refers to a situation (using Illumina as an example), where the different copies of a DNA fragment within a cluster get out of synch. If a cluster has 10,000 copies of the DNA fragment, these copies all incorporate a fluorescent nucleotide which is imaged. This nucleotide is then "de-blocked" to allow the next nucleotide to be incorporated. If the deblocking doesn't occur on a particular strand, then that strand won't incorporate the next nucleotide. But it might be deblocked in the next round, in which case it will be one base behind the rest of the cluster. As the rounds of incorporation and deblocking progress, more and more of the strands in the cluster get out of synch. The sequences might look like:

    Code:
    CATGTA
    CATGTA
    C ATGT
    CAT GT
    At this point, the imaging of the cluster will result in a mix of A and T, so the quality will be low for the basecall.
    Providing nextRAD genotyping and PacBio sequencing services. http://snpsaurus.com

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