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Guiding the design of synthetic DNA-binding molecules with massively parallel sequencing.


ABSTRACT: Genomic applications of DNA-binding molecules require an unbiased knowledge of their high affinity sites. We report the high-throughput analysis of pyrrole-imidazole polyamide DNA-binding specificity in a 10(12)-member DNA sequence library using affinity purification coupled with massively parallel sequencing. We find that even within this broad context, the canonical pairing rules are remarkably predictive of polyamide DNA-binding specificity. However, this approach also allows identification of unanticipated high affinity DNA-binding sites in the reverse orientation for polyamides containing ?/Im pairs. These insights allow the redesign of hairpin polyamides with different turn units capable of distinguishing 5'-WCGCGW-3' from 5'-WGCGCW-3'. Overall, this study displays the power of high-throughput methods to aid the optimal targeting of sequence-specific minor groove binding molecules, an essential underpinning for biological and nanotechnological applications.

SUBMITTER: Meier JL 

PROVIDER: S-EPMC3483022 | biostudies-literature | 2012 Oct

REPOSITORIES: biostudies-literature

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Guiding the design of synthetic DNA-binding molecules with massively parallel sequencing.

Meier Jordan L JL   Yu Abigail S AS   Korf Ian I   Segal David J DJ   Dervan Peter B PB  

Journal of the American Chemical Society 20121010 42


Genomic applications of DNA-binding molecules require an unbiased knowledge of their high affinity sites. We report the high-throughput analysis of pyrrole-imidazole polyamide DNA-binding specificity in a 10(12)-member DNA sequence library using affinity purification coupled with massively parallel sequencing. We find that even within this broad context, the canonical pairing rules are remarkably predictive of polyamide DNA-binding specificity. However, this approach also allows identification o  ...[more]

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