Construction of Linker-Scanning Mutations by Oligonucleotide Ligation
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The purpose of linker-scanning mutagenesis is to create a series of mutant molecules in which individual sections are sequentially replaced with the same “neutral” mutant sequence (see Fig. 1 ). Typically the technique has been applied to DNA and has most commonly been used to determine the cis -regulatory roles of upstream flanking regions of genes. The technique has several advantages over more random mutational analysis:
| 1. | The approach is systematic and implies no previous prejudice about the location of functional elements; |
| 2. | The same mutated segment is substituted in each mutant, so that potential contributions to activity from the mutated segment are minimized when comparing mutants; and |
| 3. | Most importantly, the spacing and torsional orientation of remaining functional elements remains unchanged in the mutants. |
