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  • High concentration Covaris Shearing

    Hi all,

    I'm currently using the Covaris S2 to fragment my DNA to approximately 1.5-2kb.
    Mainly so I can ligate to perform an inverse PCR before NGS library prep. My problem seems to be when I need to analyse the fragment pattern on a gel. As I only have 130ul I struggle to add enough to visualise using ethidium bromide.
    is it possible to fragment a higher concentration, i've read its independent but wondered if sonicating for a longer time would have a detrimental effect on my DNA.

    Are there any other ways round this??


    Many thanks
    Adam

  • #2
    Hi Adam,

    I can think of a couple of ways:

    (1) Use a more sensitive stain for your agarose gel. Eg, SYBR-gold.
    (2) Run a sheared DNA aliquot on an Agilent High Sensitivity chip -- but you would probably need to dilute it at least 1:5 to prevent massively overloading it. Also, you only get a picture below 10 kb or so. If you happen to have left some of the genomic DNA un-sheared, you would not see it. (But the person with their sample in the next well might...
    (3) Take the 30 ul and do a quick butanol extraction on it to reduce its volume down to something you can load in on your agarose EtBr gel.

    --
    Phillip

    Comment


    • #3
      Thats great thanks!

      Comment

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