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BMC Cancer. 2011 Jul 21;11:305. doi: 10.1186/1471-2407-11-305.

High resolution melting analysis for a rapid identification of heterozygous and homozygous sequence changes in the MUTYH gene.

Author information

1
Department of Clinical Pathophysiology, Medical Genetics Unit, University of Florence, Florence, Italy.

Abstract

BACKGROUND:

MUTYH-associated polyposis (MAP) is an autosomal recessive form of intestinal polyposis predisposing to colorectal carcinoma. High resolution melting analysis (HRMA) is a mutation scanning method that allows detection of heterozygous sequence changes with high sensitivity, whereas homozygosity for a nucleotide change may not lead to significant curve shape or melting temperature changes compared to homozygous wild-type samples. Therefore, HRMA has been mainly applied to the detection of mutations associated with autosomal dominant or X-linked disorders, while applications to autosomal recessive conditions are less common.

METHODS:

MUTYH coding sequence and UTRs were analyzed by both HRMA and sequencing on 88 leukocyte genomic DNA samples. Twenty-six samples were also examined by SSCP. Experiments were performed both with and without mixing the test samples with wild-type DNA.

RESULTS:

The results show that all MUTYH sequence variations, including G > C and A > T homozygous changes, can be reliably identified by HRMA when a condition of artificial heterozygosity is created by mixing test and reference DNA. HRMA had a sensitivity comparable to sequencing and higher than SSCP.

CONCLUSIONS:

The availability of a rapid and inexpensive method for the identification of MUTYH sequence variants is relevant for the diagnosis of colorectal cancer susceptibility, since the MAP phenotype is highly variable.

PMID:
21777424
PMCID:
PMC3156810
DOI:
10.1186/1471-2407-11-305
[Indexed for MEDLINE]
Free PMC Article

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