High-specificity DNA cleavage agent: Design and application to kilobase and megabase DNA substrates

P. Shannon Pendergrast; Yon W. Ebright; Richard H. Ebright

High-specificity DNA cleavage agent: Design and application to kilobase and megabase DNA substrates

P. Shannon Pendergrast
, Yon W. Ebright
, Richard H. Ebright

Rutgers, The State University

Research output: Contribution to journal › Article › peer-review

48 Scopus citations

Abstract

Strategies to cleave double-stranded DNA at specific DNA sites longer than those of restriction endonucleases (longer than 8 base pairs) have applications in chromosome mapping, chromosome cloning, and chromosome sequencing-provided that the strategies yield high DNA-cleavage efficiency and high DNA-cleavage specificity. In this report, the DNA-cleaving moiety copper:o-phenanthroline was attached to the sequence-specific DNA binding protein catabolite activator protein (CAP) at an amino acid that, because of a difference in DNA bending, is close to DNA in the specific CAP-DNA complex but is not close to DNA in the nonspecific CAP-DNA complex. The resulting CAP derivative, OP²⁶CAP, cleaved kilobase and megabase DNA substrates at a 22-base pair consensus DNA site with high efficiency and exhibited no detectable nonspecific DNA-cleavage activity.

Original language	American English
Pages (from-to)	959-962
Number of pages	4
Journal	Science
Volume	265
Issue number	5174
State	Published - Aug 12 1994

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abstract = "Strategies to cleave double-stranded DNA at specific DNA sites longer than those of restriction endonucleases (longer than 8 base pairs) have applications in chromosome mapping, chromosome cloning, and chromosome sequencing-provided that the strategies yield high DNA-cleavage efficiency and high DNA-cleavage specificity. In this report, the DNA-cleaving moiety copper:o-phenanthroline was attached to the sequence-specific DNA binding protein catabolite activator protein (CAP) at an amino acid that, because of a difference in DNA bending, is close to DNA in the specific CAP-DNA complex but is not close to DNA in the nonspecific CAP-DNA complex. The resulting CAP derivative, OP26CAP, cleaved kilobase and megabase DNA substrates at a 22-base pair consensus DNA site with high efficiency and exhibited no detectable nonspecific DNA-cleavage activity.",

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AB - Strategies to cleave double-stranded DNA at specific DNA sites longer than those of restriction endonucleases (longer than 8 base pairs) have applications in chromosome mapping, chromosome cloning, and chromosome sequencing-provided that the strategies yield high DNA-cleavage efficiency and high DNA-cleavage specificity. In this report, the DNA-cleaving moiety copper:o-phenanthroline was attached to the sequence-specific DNA binding protein catabolite activator protein (CAP) at an amino acid that, because of a difference in DNA bending, is close to DNA in the specific CAP-DNA complex but is not close to DNA in the nonspecific CAP-DNA complex. The resulting CAP derivative, OP26CAP, cleaved kilobase and megabase DNA substrates at a 22-base pair consensus DNA site with high efficiency and exhibited no detectable nonspecific DNA-cleavage activity.

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High-specificity DNA cleavage agent: Design and application to kilobase and megabase DNA substrates

Abstract

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