Browsing by Subject "amplifications"
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Item Analysis of Low Copy Number DNA Using Profiler Plus at Increased Amplification Cycles and Modifications in Sample Injection Parameters(2003-08-01) Hynds, Jody Lynn; Arthur Eisenberg; John Planz; Joseph WarrenThere are many DNA testing techniques that can be utilized for samples with low quantities of DNA. Mitochondrial DNA testing is designed for successful DNA sequencing of hair shafts, degraded and burned samples. Newly developed SNP (single nucleotide polymorphisms) testing is also designed for the analysis of challenging samples. The increased interest in the analysis of low copy number DNA samples using STR testing is necessitated since the national database CODIS (Combined Data Index System) currently only accepts the DNA profiles analyzed with the 13 core STR loci. CODIS contains DNA profiles of evidence found at crime scenes, convicted offender and missing persons DNA profiles (4). The goal of this project is to develop methodologies to increase the success rate of LCN DNA samples using STR testing. The experimental design for this study involved the amplification of DNA isolated from buccal swabs using the Profiler Plus multiplex kit at two different DNA input quantities: 0/0156ng (15.6pg) and 0.0312ng (31.2pg). Four separate amplifications of these DNA samples were done at: 28, 30, 32 and 34 cycles. The manufacturer’s recommended cycle number for AmpFISTR Profiler Plus is 28 cycles. These samples were analyzed on both the ABI Prism 310 Genetic Analyzer and the ABI Prism 3100 Genetic Analyzer using OCD standard protocols for loading samples. The injection time and voltage were modified for each of the number of PCR cycles. The best combination of cycle number and injection parameters was chosen for the low copy number reproducibility study.Item Evaluation or the Gold-Plated Silver Sample Block on the GeneAmp PCR System 9700 Thermal Cycler(2003-05-01) Moore, Melody Ann; Arthur Eisenberg; Joseph Warren; John PlanzMoore, Melody Ann. Evaluation of the Gold-plated Silver Sample Block on the GeneAmp PCR System 9700 Thermal Cycler. Master of Science (Forensic Genetics), May 2003, 49 pages, 9 figures, 3 tables, 2 appendices, 24 references. The GeneAmp PRC System 9700 Thermal Cycler has been introduced with interchangeable silver and gold-plated silver sample blocks. To validate the new gold-plated silver sample block on the System 9700, amplifications were performed on both the gold-plated silver and the previously validated silver sample blocks. PCR amplifications using the AmpFLSTR Profiler Plus ID COfiler, Identifiler and SGM Plus typing kits (Applied Biosystems, Forster City, CA) were performed. Electrophoretic characteristics such as allele concordance, peak heights, and peak height ratios were used to discern any differences in the amplification capabilities of the two sample blocks. The results demonstrate that the PCR reactions on both silver and gold-plated silver blocks are equivalent. In addition, the data obtained (allele cells, peak heights, peak height ratios) from both Macintosh and Windows NT platforms were identical for each amplification kit, indicating concordance between the software packages of each operating system. The results of this study demonstrate the interchangeability of the silver and gold-plated sample blocks.