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Development of Novel Diagnostics for Fragile X Syndrome

脆性 X 综合征新型诊断方法的开发

基本信息

批准号：
7908031
负责人：
SEIYU HOSONO
金额：
$ 53.27万
依托单位：
JS GENETICS, INC.
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-04-25 至 2012-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7908031
关键词：
5&apos Untranslated Regions Adult Autistic Disorder Award Base Pairing Binding Biological Assay Blood Blood capillaries Boston CGG repeat CGG repeat expansion Child Chromosome Band Clinical DNA Development Diagnosis Diagnostic Diagnostic tests Disease Electrophoresis Etiology FMR1 FMR1 Gene FXTAS Family history of Female Fetus Fragile X Gene Fragile X Mental Retardation Protein Fragile X Syndrome Future Generations Gel Gene Mutation Genes Genetic Genetic screening method Genome Guanosine Guanosine Triphosphate Incidence Individual Infertility Laboratories Language Lead Learning Disabilities Mental Retardation Methods Methylation Molecular Molecular Weight Motor Neonatal Screening Neurons Parkinsonian Disorders Pediatric Hospitals Pervasive Development Disorder Phase Primary Health Care Professional counselor Promoter Regions Proteins Provider Reaction Risk Sampling Screening procedure Site Small Business Innovation Research Grant Southern Blotting Speech Spottings Stretching Syndrome Taq Polymerase Testing Time Translating Translations Tremor Trinucleotide Repeats Untranslated Regions Woman X Chromosome base capillary commercial application cost design equilibration disorder gel electrophoresis high throughput screening male medical schools meetings neurodevelopment novel novel diagnostics nucleotide analog pregnant prenatal testing programs public health relevance rapid technique tool

项目摘要

DESCRIPTION (provided by applicant): Fragile X Syndrome (FRAX) is the most common genetic cause of mental retardation in males. FRAX is caused by the expansion of a CGG trinucleotide repeat of the 5' untranslated region (UTR) of the FMR1 gene. In normal individuals, the 5' UTR of the FMR1 gene contains 5 to 45 CGG repeats; however, individuals with FRAX have over 200 repeats. Presently, Southern Blot analysis is used to determine the size of the repeat segment and methylation status of the FRAX gene. This test only detects the gross size of CGG repeats and is labor intensive and expensive. PCR and gel electrophoresis is typically used to determine the size of the CGG expansion. This approach is limited, as PCR reactions typically fail to amplify long stretches of CGG expansions (>25 repeats) and molecular weight determination by electrophoresis via capillary or slab gels is labor intensive. In our Phase I SBIR award, we developed a novel, highly efficient and accurate screening test for diagnosing FRAX. In Step 1, Whole Genome Multiple Displacement Amplification using 7-deaza-2-Guanosine (7-deaza GTP) nucleotide analog is incorporated into multiple copies of the CGG FMR1 expansion. In Step 2, Site Specific Multiple Displacement Amplification (SSMDA) using 7-deaza GTP is performed to weaken the GC base pairings, making the GCC expansion more accessible to Taq DNA Polymerase in real-time PCR. In Step 3, SSMDA is followed by quantitative assessment of the numbers of CGG triplet repeats using TaqMan real-time PCR without the need for sizing by gel electrophoresis or Southern blotting. We hypothesize that using this new molecular-based method, we can develop an effective, rapid and low-cost screening test for FRAX with broad commercial application. In this Phase II application we propose to: SA1: Optimize our assay for CGG repeat copy number determination using a large number of DNA samples with varying degrees of CGG repeats. SA2: Test our assay in a clinical FRAX diagnostic laboratory. SA3: Test our assay in blood spots for potential newborn screening. We anticipate that this Phase II application will lead to the optimization and implementation of a test that is suitable for low cost high-throughput screening for FRAX. As such, this test has the potential to markedly change how we currently screen for FRAX. These studies will involve the combined expertise of JS Genetics, which has been developing novel DNA-based diagnostic tests, and Dr. Bai-Lin Wu of Boston Children's Hospital and Harvard Medical School, who an extensive track record of developing clinical DNA diagnostic studies and in clinical laboratory testing of FRAX. PUBLIC HEALTH RELEVANCE: Fragile X Syndrome (FRAX) is the most common genetic cause of mental retardation in males. We propose the development of a new molecular-based method, for rapid and low-cost screening of FRAX with broad commercial application.

描述（由申请人提供）：