Method to Detect Mutations in Human Ataxia-Telangiectasia Mutation (ATM gene)
Ataxia-Telangiectasia (AT) is an autosomal recessive genetic disorder caused by mutation in ATM (AT Mutation) gene. It is a primary immunodeficiency disease with an early onset and is generally fatal with in the first two decades of life. The individuals afflicted with AT have 100 fold increased risk of developing cancer, with estimated 10% of AT patients developing lymphoma or leukemia. In the United States as many as 2.5 million people may carry a single copy of the mutated ATM gene. The carrier state for the defective ATM is linked to breast cancer with up to 8% of the breast cancer cases being associated with it. Additionally, the defects in ATM gene result in increased sensitivity to iodizing radiation and therefore increased risk of radiation burns due to chemotherapy.
The researchers at Benaroya Research Institute have developed a method to detect mutations in human ATM gene using novel sets of PCR primers. The primers are designed to amplify genomic DNA corresponding to each of the 65 coding exons and the flanking sequences to ensure detection of splice site mutations. To detect the mutations, DNA sample from 1) a subject suspected to have mutation(s) and 2) a control sample without mutation, are PCR amplified using the primers. The sequence differences may be detected using methods such as DNA sequencing or gel electrophoresis. Subset of primers can be used together in a PCR reaction for a simultaneous multiplex analysis of two or more exons. The multiplex PCR products may be analyzed using an electrophoresis method, such as single stranded conformation polymorphism (SSCP).
Over 200,000 new cases of breast cancer are diagnosed annually in the United States, suggesting that over 15,000 of those cases may be attributed to defect in ATM gene. The increased radiation sensitivity due to the ATM mutation also increases the cancer risk due to workplace radiation exposure of the carriers. Detection of the ATM mutation is a critical step in minimizing the cancer risk for healthy carriers and preventing unwanted radiation damage in cancer patient carrying ATM mutation.
The researchers have successfully identified 30 mutations in ATM gene using the designed primer sets. The multiplex analysis using SSCP as the detection method was successful for the defined subsets of primers.
Issued US Patent 5,770,372: Detection of Mutations in The Human ATM Gene, 06/23/1998
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