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The effect of UDG treatment on Kunkel and selective rolling circle amplification (RCA) mutagenesis.

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posted on 20.02.2013, 09:03 by Tuomas Huovinen, Eeva-Christine Brockmann, Sultana Akter, Susan Perez-Gamarra, Jani Ylä-Pelto, Yuan Liu, Urpo Lamminmäki

CDR-H3 region was altered by Kunkel mutagenesis and further treated with +/− UDG and amplified with RCA. The mutant yield was studied by plating transformed samples. Successful CDR-H3 primer incorporation in scFv-β-lactamase gene resulted in ampicillin resistant clones (pie chart, black sector). The template clones were sensitive to ampicillin (pie chart, white sector). In theory, 22% of the mutated clones with 7× NNN codons are not ampicillin resistant due to primer-born STOP-codons (pie chart, grey sector).

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