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Mutat Res. 2009 Jun-Jul;677(1-2):27-32. doi: 10.1016/j.mrgentox.2009.05.003. Epub 2009 May 13.

Detection of K-ras mutations in azoxymethane-induced aberrant crypt foci in mice using LNA-mediated real-time PCR clamping and mutant-specific probes.

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Department of Medicine, Flinders Centre for Cancer Prevention and Control, Flinders University of South Australia, Adelaide, Australia.


Azoxymethane, a rodent colon-specific carcinogen, induce DNA damage, and causes proto-oncogene K-ras point mutations and subsequent tumor formation if DNA damage is not repaired or removed. The present study was designed to detect and characterize K-ras mutations in azoxymethane (AOM)-induced aberrant crypt foci (ACF) in mice, and determine whether dietary supplementation of selenium influences K-ras mutations frequency in ACF using a new PCR technique of locked nucleic acid-mediated real-time PCR clamping combined with mutant-specific probes. K-ras mutations were identified in 33% of AOM-induced ACF. In addition to G to A transition mutation, specific G to T transversion mutation was also identified for the first time in mouse ACF. Furthermore, selenium intake was associated with reduced ACF formation and reduced K-ras mutations rate, respectively, from 112 and 37% in mice fed control diet to 65 and 14% in mice fed selenium-containing diet (p < 0.05). This is the first report of the use of one-step LNA-mediated real-time PCR clamping to detect K-ras mutations in AOM-induced colon cancer model. It is highly sensitive and can be applied to the detection of early genetic alterations in carcinogen-based animal models.

[Indexed for MEDLINE]

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