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LigAmp for sensitive detection of single-nucleotide differences.

Publication ,  Journal Article
Shi, C; Eshleman, SH; Jones, D; Fukushima, N; Hua, L; Parker, AR; Yeo, CJ; Hruban, RH; Goggins, MG; Eshleman, JR
Published in: Nat Methods
November 2004

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.

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Published In

Nat Methods

DOI

ISSN

1548-7091

Publication Date

November 2004

Volume

1

Issue

2

Start / End Page

141 / 147

Location

United States

Related Subject Headings

  • ras Proteins
  • Sequence Analysis, DNA
  • Sequence Alignment
  • Sensitivity and Specificity
  • Reverse Transcriptase Polymerase Chain Reaction
  • Reproducibility of Results
  • Proto-Oncogene Proteins p21(ras)
  • Proto-Oncogene Proteins
  • Polymorphism, Single Nucleotide
  • Pancreatic Neoplasms
 

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Shi, C., Eshleman, S. H., Jones, D., Fukushima, N., Hua, L., Parker, A. R., … Eshleman, J. R. (2004). LigAmp for sensitive detection of single-nucleotide differences. Nat Methods, 1(2), 141–147. https://doi.org/10.1038/nmeth713
Shi, Chanjuan, Susan H. Eshleman, Dana Jones, Noriyoshi Fukushima, Li Hua, Antony R. Parker, Charles J. Yeo, Ralph H. Hruban, Michael G. Goggins, and James R. Eshleman. “LigAmp for sensitive detection of single-nucleotide differences.Nat Methods 1, no. 2 (November 2004): 141–47. https://doi.org/10.1038/nmeth713.
Shi C, Eshleman SH, Jones D, Fukushima N, Hua L, Parker AR, et al. LigAmp for sensitive detection of single-nucleotide differences. Nat Methods. 2004 Nov;1(2):141–7.
Shi, Chanjuan, et al. “LigAmp for sensitive detection of single-nucleotide differences.Nat Methods, vol. 1, no. 2, Nov. 2004, pp. 141–47. Pubmed, doi:10.1038/nmeth713.
Shi C, Eshleman SH, Jones D, Fukushima N, Hua L, Parker AR, Yeo CJ, Hruban RH, Goggins MG, Eshleman JR. LigAmp for sensitive detection of single-nucleotide differences. Nat Methods. 2004 Nov;1(2):141–147.

Published In

Nat Methods

DOI

ISSN

1548-7091

Publication Date

November 2004

Volume

1

Issue

2

Start / End Page

141 / 147

Location

United States

Related Subject Headings

  • ras Proteins
  • Sequence Analysis, DNA
  • Sequence Alignment
  • Sensitivity and Specificity
  • Reverse Transcriptase Polymerase Chain Reaction
  • Reproducibility of Results
  • Proto-Oncogene Proteins p21(ras)
  • Proto-Oncogene Proteins
  • Polymorphism, Single Nucleotide
  • Pancreatic Neoplasms