Cloning and Characterization of Expanded CAG-Repeat Containing Sequence(s): Identification of Candidate Breast Cancer Predisposition Gene(s)

摘要

Trinucleotide repeats (TNRs) are widely present in the human genome, and their expansions have been recognized to he the cause of several genetic disorders. In a previous study, we have identified expanded (CAG) repeats in 2.4% of breast cancer cases (n=212). No expansion of the sane magnitude has been detected in 196 population controls. In the current project our objective is to identify and characterize such sequences in breast cancer specimens. We have developed an effective cloning and screening strategy using a combination of dynabead enrichment microsatellite isolation protocol (Glenn-Schable protocol). Colonies were selected and sequenced to identify CAG repeat containing fragments from breast cancer patients. We have shown that 20% of the colonies contained polymorphic (larger/smaller) repeat lengths compared to corresponding sequences deposited at NCBI. Such sequences resemble polymorphic CAG repeats that may influence the intrinsic properties and function of nearby genes. To complement the library construction, we have also systematically identified CAG repeat containing coding sequences from cancer genes using Bioinformatics. In this study we have implicated the presence of polymorphic CAG repeats within the functional domains of cancer proteins. Molecular cloning and bioinformatics approaches used in this study has provided a valuable resource for polymorphic CAG-containing sequences within relevant cancer genes. Future direction involves genetic and epidemiological studies, where the risk contributed by CAG repeat expansions to breast cancer risk will be evaluated.