• Directed mismatch
  • Directed mismatch allows you to create or remove restriction sites on the current DNA sequence or its variants by incorporating single or double mismatches at a site near the mutation. Using this function you can create or destroy restriction sites in order to distinguish the wild type allele from a common mutant allele.

    1.Choose the Restriction | Directed Mismatch command to open the Directed Mismatch dialog box.

    2.Select a restriction enzyme file and the Cutter option. “restrict.enz” is the default enzyme file in DNAMAN.

    3.Type a number in the Mutation at box to select the mutant position.

    4.Type A, C, G or T code in the for box to define the mutation.

    6.Click the Site button to confirm the sequence and mutation.

    7.Select one of the Accept Mutation options:

    Non option searches for restriction sites without any mismatch.

    <= 1” option searches for restriction sites that can accept a single mismatch.

    <= 2 option searches for restriction sites that can accept double mismatches.

    Example:

    1      TTTGACTGCCACTTCCTCGATGAAGGTTTTACTGCCAAGG
           -------------------C--------------------

    Mutation at position 20

    Maximum Mismatch = 2

    W.T. indicates the wild type sequence

    '*' indicates mutation position

    '^' indicates mismatch

    AflII

    Mutant:CTGAAG

    Enzyme:CTTAAG

        * ^   

    A single mismatch “T” near the mutant allele “C” creates the enzyme site of AflII in the mutant sequence, but not in the wild type sequence.

    Eco57I 
    Mutant:    CTGAAG
    Enzyme:    CTGAAG
            *

    The enzyme site of Eco57I exists in the mutant without incorporating any mismatch.

    XbaI 
    W.T. :  CCTCGA
    Enzyme: TCTAGA
            ^  ^ *

    Double mismatches “T” and “A” near the wild type allele “A” creates the enzyme site of XbaI in the wild type sequence, but not in the mutant.