Tagging DNA mismatches by selective 2'-amine acylation.
BACKGROUND
Widespread characterization of genetic variation and disease at the gene-sequence level has inaugurated a new era in human biology. Techniques for the molecular analysis of these variations and their linkage with measurable phenotypes will profoundly affect diverse fields of biological chemistry and biology.


RESULTS
A chemical tagging method has been developed to detect point mutations and other defects in nucleic acid sequences. The method employs oligodeoxynucleotide probes in which one 2'-ribose position (-H) is substituted with an amine (-NH(2)) group. 2'-Amine-substituted nucleotides are specifically acylated by succinimidyl esters to form a 2'-amide product. The mutation detection method exploits our observation that 2'-amine groups at the site of a mismatch are acylated more rapidly than amine substitutions at base-paired nucleotides. 2'-Amine acylation is governed primarily by local, rather than global, differences in nucleotide dynamics, such that site-specific tagging of DNA mismatches does not require discriminatory hybridization conditions to be determined.


CONCLUSIONS
2'-Amine mismatch tagging offers an approach for chemically interrogating the base-paired state of individual nucleotides in a hybridized duplex and for quantifying nucleicacid hybridization with single-base specificity.
John
D M
DM
Department of Chemistry, University of North Carolina, NC 27599-3290, USA.
Weeks
K M
KM
D000215:Acylation; D020137:Base Pair Mismatch; D004247:DNA / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D009693:Nucleic Acid Hybridization; D013816:Thermodynamics
D004247:DNA