Genetic disease detection and DNA amplification using cloned thermostable ligase Academic Article Article uri icon

Overview

MeSH Major

  • Neoplasms
  • Polymorphism, Single Nucleotide

abstract

  • Polymerase chain reaction, using thermostable DNA polymerase, has revolutionized DNA diagnostics. Another thermostable enzyme, DNA ligase, is harnessed in the assay reported here that both amplifies DNA and discriminates a single-base substitution. This cloned enzyme specifically links two adjacent oligonucleotides when hybridized at 65 degrees C to a complementary target only when the nucleotides are perfectly base-paired at the junction. Oligonucleotide products are exponentially amplified by thermal cycling of the ligation reaction in the presence of a second set of adjacent oligonucleotides, complementary to the first set and the target. A single-base mismatch prevents ligation/amplification and is thus distinguished. This method was exploited to detect 200 target molecules as well as to discriminate between normal beta A- and sickle beta S- globin genotypes from 10-microliters blood samples.

publication date

  • January 17, 1991

Research

keywords

  • Academic Article

Identity

Digital Object Identifier (DOI)

  • 10.1073/pnas.88.1.189

PubMed ID

  • 1986365

Additional Document Info

start page

  • 189

end page

  • 93

volume

  • 88

number

  • 1