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Replicates of Rec-seq experiments with wild-kind Cre cluster together toward the middle of the graph; Ala-substituted mutants that improve sequence desire appear to the left of the wild-sort grouping, while desire-diminishing variants cluster to the proper. Structural and mutagenesis studies23,26,31 urged that mutation of Arg259 would affect specificity at half-site position 10. Indeed, the Arg259→Ala variant confirmed a drop in enrichment at place 10 (from 5.0-fold for wild-type Cre to 1.1-fold for the mutant), with a modest preference for C or T in the left half-site and G or A in the right half-site (Fig. 2b, c). Rec-seq revealed the sequence choice of Cre to be asymmetric, as is obvious when the left and proper half-site enrichment profiles are superimposed (Supplementary Fig. 6a). To ensure that an asymmetric sequence choice is a property of the enzyme and never as a result of totally different DNA sequences flanking the library oligonucleotides (Fig. 1b), we carried out Rec-seq utilizing a substrate library an identical to the unique except that the non-palindromic loxP core was replaced with its reverse complement (Supplementary Fig. 6b). The Rec-seq enrichment profile of this "inverted core" loxP library mirrored, reasonably than duplicated, the profile on the unique substrate library (Supplementary Fig. 6b), indicating that the oligonucleotide sequence context was not answerable for the asymmetry of the Cre specificity profile

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