Detection of colorectal cancer k-ras mutations using a simplified oligonucleotide ligation assay

Cynthia B. Rothschild, Catherine S. Brewer, Brian W. Loggie, G. Alan Beard, Mark X. Triscott

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Abstract

It has been suggested that some mutations in codons 12 and 13 of the K- ras gene are associated with the progression of colorectal adenomas to carcinomas. The aim of this study was to develop a rapid, colorimetric assay for K-ras point mutations commonly associated with colorectal cancer. K-ras exon 1 was amplified from colorectal tumor DNA and K-ras activating mutations detected using an oligonucleotide ligation assay (OLA) in combination with immunological and colorimetric detection. Using the OLA with oligonucleotides specific to individual K-ras mutations, 6 (of 17 total colorectal adenomas/carcinomas) were found to have K-ras mutations. The assay could detect as little as 10% mutant allele. A simplified OLA designed to test for either the presence (+) or absence (-) of any of the K-ras activating mutations was developed. The assay was further streamlined by use of a dipstick methodology for colorimetric development. If required, assay sensitivity can be increased by the use of the recently described EDNA-ELCA detection system. The simplified (+/-) mutation OLA in combination with a dipstick or EDNA-ELCA detection system provides a rapid, sensitive assay for K-ras point mutations suitable for use as part of the clinical assessment of colorectal cancer.

Original languageEnglish
Pages (from-to)11-19
Number of pages9
JournalJournal of Immunological Methods
Volume206
Issue number1-2
DOIs
Publication statusPublished - Aug 7 1997
Externally publishedYes

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All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Immunology

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Detection of colorectal cancer k-ras mutations using a simplified oligonucleotide ligation assay. / Rothschild, Cynthia B.; Brewer, Catherine S.; Loggie, Brian W.; Beard, G. Alan; Triscott, Mark X.

In: Journal of Immunological Methods, Vol. 206, No. 1-2, 07.08.1997, p. 11-19.

Research output: Contribution to journalArticle