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  • Nextera XT library preparation

    I am using Nextera XT for library preparation.
    The kit suggests 12 cycles of PCR after tagmentation.
    If I run 5 ul of PCR product after 12 cycles I do not see any smear on my 1.5% agarose gel. If I run 13 cycles then I see the smear.
    I am wondering if this is normal or if I should see a smear after 12 cycles.
    I will try to go to the Bioanalyzer to see my library but I am not sure it will give me a better answer.
    Any help or comment could be really useful.

  • #2
    I do not run my libraries on gel to check, so I do not know if one should see smear or not. If you see smear after 13 cycles by running 5 ul of PCR, then using 10 ul after 12 cycles should give similar results.

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    • #3
      This is normal. Many people modify the protocol to increase the number of PCR cycles like this.

      Comment

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