BOMB protocol #6.5 TNA extraction from yeast

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  • #1591
    PStepper
    Keymaster

      reserved space

      #18220
      Iason K.
      Participant

        Hi ,

        I have been using the bomb bio 6.5 protocol for yeast and works like a charm. It provide us with  long DNA fragments that we require but I want mainly the gDNA. Can I use RNase A and at which step would you recommend and at what concentration?

        #18237
        admin
        Keymaster

          Ahoy,

          I am stocked to hear that you like our protocol!
          I presume that you have thought through whether RNA contamination actually poses an issue.
          Because unless you are doing super sensitive quantification, transfections or steps involving reverse transcriptase, I never really encounter any issues whatsoever with residual RNA. But, I am mostly  running PCRs or similarly robust techniques.

          I am not sure whether RNAse A has a tolerance for Lyticase and GITC but I would not risk it.

          Unfortunately the save option would be to finish the protocol transfer your elution to a tube loaded with RNAse A + Buffer –> Incubate  (+ maybe deactivate) and then perform something like a PEG (SPRI \ Ampure) purification or if you want to go DIY all the way BOMB protocol 4.1., obviously you can also ethanol precipitate etc at this point.

          Cheers

          Tim M

           

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