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Genetics. 2019 Jul;212(3):655-665. doi: 10.1534/genetics.119.302268. Epub 2019 May 24.

Quantifying Microsatellite Mutation Rates from Intestinal Stem Cell Dynamics in Msh2-Deficient Murine Epithelium.

Author information

1
Li Ka Shing Centre, Cancer Research UK Cambridge Institute, University of Cambridge, CB2 0RE, United Kingdom.
2
Department Biological and Medical Sciences, Faculty of Health and Life Sciences, Oxford Brookes University, OX3 0BP, United Kingdom.
3
MRC Weatherall Institute of Molecular Medicine, University of Oxford, OX3 9DS, United Kingdom.
4
Li Ka Shing Centre, Cancer Research UK Cambridge Institute, University of Cambridge, CB2 0RE, United Kingdom doug.winton@cruk.cam.ac.uk.

Abstract

Microsatellite sequences have an enhanced susceptibility to mutation, and can act as sentinels indicating elevated mutation rates and increased risk of cancer. The probability of mutant fixation within the intestinal epithelium is dictated by a combination of stem cell dynamics and mutation rate. Here, we exploit this relationship to infer microsatellite mutation rates. First a sensitive, multiplexed, and quantitative method for detecting somatic changes in microsatellite length was developed that allowed the parallel detection of mutant [CA]n sequences from hundreds of low-input tissue samples at up to 14 loci. The method was applied to colonic crypts in Mus musculus, and enabled detection of mutant subclones down to 20% of the cellularity of the crypt (∼50 of 250 cells). By quantifying age-related increases in clone frequencies for multiple loci, microsatellite mutation rates in wild-type and Msh2-deficient epithelium were established. An average 388-fold increase in mutation per mitosis rate was observed in Msh2-deficient epithelium (2.4 × 10-2) compared to wild-type epithelium (6.2 × 10-5).

KEYWORDS:

Msh2; intestine; microsatellite; mouse; mutation

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