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Microarray based STR genotyping utilizing RecA-mediated ligation

利用 RecA 介导的连接进行基于微阵列的 STR 基因分型

基本信息

批准号：
8098886
负责人：
ROBERT E WAGNER
金额：
$ 38.74万
依托单位：
GENE CHECK, 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：
2008-05-01 至 2012-09-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8098886
关键词：
Alleles Base Pairing Biological Assay Blood Code Collection Computer software DNA DNA biosynthesis Data Detection Development Diagnosis Disease Exhibits Filament Forensic Medicine Fractionation Generations Genes Genetic Genetic Markers Genetic Polymorphism Genetic Predisposition to Disease Genomics Genotype Goals Hereditary Disease Histocompatibility Testing Hour Human Human Genome Human Identifications Human Resources Individual Institutes Investigation Laboratories Lead Length Ligation Livestock Mediating Medical Methods Microsatellite Repeats Oligonucleotide Probes Oligonucleotides Performance Phase Plants Police Probability Process Protocols documentation Reagent Relative (related person)Resources Sales Sampling Scheme Signal Transduction Site Slide Solutions Specificity Speed System Systems Analysis Tandem Repeat Sequences Technology Testing Time Trinucleotide Repeats Whole Blood Wild Animals Work base commercialization companion animal cost cost effective ds-DNA father role genetic analysis member novel public health relevance research study success

项目摘要

DESCRIPTION (provided by applicant): Tandem repeat sequences (STR) are distributed widely in the human genome. These sites exhibit high levels of length polymorphism as a result of their relative instability during DNA replication, i.e., the tendency for slippage to occur between the template and newly synthesized strands causing deletion or addition of repeat units. Changes in STR repeat number can lead to heritable genetic predispositions and disease. However, their variability makes them ideal markers for genetic studies. One class, the microsatellites containing 4 base pair repeat units, has been selected by the FBI as the basis for a genetic identification system, termed CODIS. Current analysis systems require a 1-2 day assay and employ electrophoretic separation of DNA fragments that contain the STR locus for accurate sizing and identification of the repeat number. The long-term goal of this project is to develop a simple, cost-effective and automatable assay system for STR analysis that allows human identification in less than 3 hours from sample to result by avoiding the need for PCR amplification, target DNA denaturation and electrophoretic separation. In Phase I of this proposal a single locus (TPOX) was used to demonstrate that accurate microarray based STR analysis is technically feasible and sufficiently rapid. In Phase II we will expand this assay to include at least 6 other CODIS loci, convert the technology from an in solution to an immobilized format and attempt to increase assay speed and convenience further by eliminating the PCR step. Our approach to achieving these goals utilizes Gene Check's RecA mediated ligation (RML) technology in which the extreme precision of RecA mediated homology searching in double stranded DNA is used to catalyze specific base pairing of oligonucleotide probes at the STR locus with subsequent ligation into specific products that reflect the length of the STR locus. Gene Check has discovered that RML can operate efficiently when one of the oligonucleotides is immobilized to a microarray slide, making high order multiplexing of the RML-STR technology feasible and practical. Signal amplification methods will be employed to allow genotyping directly from double stranded genomic DNA. Success in this project will produce an STR panel that can immediately be used in forensic applications as well as in human paternity and identity testing. The RML-STR system can readily be adapted to livestock, companion animal and wild animal parentage and identity testing and has potential application to diagnosis of medical conditions related to microsattelite sequence expansion. PUBLIC HEALTH RELEVANCE: Gene Check proposes to utilize its RecA mediated ligation technology for STR genotyping (RML-STR) in a microarray format to allow robust, rapid, low cost and automated STR genotyping. This system will be useful for personnel identification in everyday police investigations as well as in a variety of other forensic and non-human identification applications. Medical applications include tissue typing and genetic analysis of triplet repeat associated disorders.

描述（由申请人提供）：