High-specificity DNA cleavage agent: design and application to kilobase and megabase DNA substrates.
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.
Pendergrast
P S
PS
Department of Chemistry, Rutgers University, New Brunswick, NJ 08855.
Ebright
Y W
YW
Ebright
R H
RH
D001483:Base Sequence; D002373:Cyclic AMP Receptor Protein / Q000737:chemistry*; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009690:Nucleic Acid Conformation; D010618:Phenanthrolines / Q000737:chemistry*
D002373:Cyclic AMP Receptor Protein; D004268:DNA-Binding Proteins; D010618:Phenanthrolines; C088677:catabolite activator protein, copper-OS26; D004247:DNA; C025205:1,10-phenanthroline