Biology Forum Molecular Biology Quick Change

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    • #1981

      I am having issues with Quick change mutagenesis. 🙁 🙁
      I am trying to intrduce a single base change in a gene cloned into a plasmid (5Kb). My oligos are 37 bp long with 18 bp homology on either side.
      I am following the protocol as per the manual but only variation is that
      a) My oligos are not PAGE purified
      b) I donot use the super competent cells for transformation. I make my own(chemically competent).
      At the end of it I get NO colonies.:shock: 😯 😯
      Is my reaction not working or my transformation?????????/
      In either case what would you recomend???????/
      PL. chip in with idea(s). Any suggession(s) wil be appreciated.

    • #29988

      At least you should have a transformation control, is it working? If not The problem is coming from your cells, if it does, from your mutagenesis, but I am not familiar with thi sprotocol, so I can’t help…

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