Overcoming problems with limiting DNA samples in forensics and clinical diagnostics using multiple displacement amplification
Muharam, Firman Alamsyah (2006) Overcoming problems with limiting DNA samples in forensics and clinical diagnostics using multiple displacement amplification. .
The availability of DNA samples that are of adequate quality and quantity is essential for any genetic analysis. The fields of forensic biology and clinical diagnostic pathology testing often suffer from limited samples that yield insufficient DNA material to allow extensive analysis. This study examined the utility of a recently introduced whole genome amplification method termed Multiple Displacement Amplification (MDA) for amplifying a variety of limited sample types that are commonly encountered in the fields of forensic biology and clinical diagnostics. The MDA reaction, which employs the highly processive bacteriophage φ29 DNA polymerase, was found to generate high molecular weight template DNA suitable for a variety of downstream applications from low copy number DNA samples down to the single genome level. MDA of single cells yielded sufficient DNA for up to 20,000,000 PCR assays, allowing further confirmatory testing on samples of limited quantities or the archiving of precious DNA material for future work. The amplification of
degraded DNA material using MDA identified a requirement for samples of sufficient quality to allow successful synthesis of product DNA templates. Furthermore, the utility of MDA products in comparative genomic hybridisation
(CGH) assays identified the presence of amplification bias. However, this bias was overcome by introducing a novel modification to the MDA protocol. Future directions for this work include investigations into the utility of MDA products in short tandem repeat (STR) assays for human identifications and application of the modified MDA protocol for testing of single cell samples for genetic abnormalities.
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|Item Type:||QUT Thesis (Masters by Research)|
|Supervisor:||van Daal, Angela& Morris, Charles|
|Keywords:||whole genome amplification, multiple displacement amplification, φ29, phi29, DNA polymerase, single cell, low copy number, DNA, PCR, forensic biology, forensic science, clinical diagnostics, comparative genomic hybridisation|
|Divisions:||Past > QUT Faculties & Divisions > Faculty of Science and Technology|
Past > Schools > School of Life Sciences
|Department:||Faculty of Science|
|Institution:||Queensland University of Technology|
|Copyright Owner:||Copyright Firman Alamsyah Muharam|
|Deposited On:||03 Dec 2008 13:58|
|Last Modified:||29 Oct 2011 05:44|
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