LigAmp for sensitive detection of single-nucleotide differences.

Published

Journal Article

We developed the LigAmp assay for sensitive detection and accurate quantification of viruses and cells with single-base mutations. In LigAmp, two oligonucleotides are hybridized adjacently to a DNA template. One oligonucleotide matches the target sequence and contains a probe sequence. If the target sequence is present, the oligonucleotides are ligated together and detected using real-time PCR. LigAmp detected KRAS2 mutant DNA at 0.01% in mixtures of different cell lines. KRAS2 mutations were also detected in pancreatic duct juice from patients with pancreatic cancer. LigAmp detected the K103N HIV-1 drug resistance mutation at 0.01% in plasmid mixtures and at approximately 0.1% in DNA amplified from plasma HIV-1. Detection in both systems is linear over a broad dynamic range. Preliminary evidence indicates that reactions can be multiplexed. This assay may find applications in the diagnosis of genetic disorders and the management of patients with cancer and infectious diseases.

Full Text

Duke Authors

Cited Authors

  • Shi, C; Eshleman, SH; Jones, D; Fukushima, N; Hua, L; Parker, AR; Yeo, CJ; Hruban, RH; Goggins, MG; Eshleman, JR

Published Date

  • November 2004

Published In

Volume / Issue

  • 1 / 2

Start / End Page

  • 141 - 147

PubMed ID

  • 15782177

Pubmed Central ID

  • 15782177

International Standard Serial Number (ISSN)

  • 1548-7091

Digital Object Identifier (DOI)

  • 10.1038/nmeth713

Language

  • eng

Conference Location

  • United States