Results from the NIST 2004 DNA Quantitation Study

Kline, Margaret C.; Duewer, David L.; Redman, Janette W.; Butler, John M.

doi:10.1520/jfs2004357

Cited by 57 publications

(43 citation statements)

References 28 publications

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“…The variation in the estimated mtGE found in the temporal reproducibility study (at worst about a factor of 2) was similar to the results we obtained with a commercially available rtPCR kit for quantification of amplifiable human nuclear DNA (data not shown) and to data from the literature [5,9].…”

Section: Temporal Reproducibilitysupporting

confidence: 86%

Characterization of mtDNA SNP typing and mixture ratio assessment with simultaneous real-time PCR quantification of both allelic states

Niederstätter

Coble²,

Grubwieser

et al. 2005

Int J Legal Med

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We performed a study on the forensic utility of allele-discriminatory quantitative real-time PCR (rtPCR) using Minor Groove Binder TaqMan probes, targeting the highly variable mitochondrial single nucleotide polymorphism 16519T/C. The apparent single-cycle PCR efficiency was virtually 100% for both 16519 alleles. The allele designations made by rtPCR were concordant with the results obtained in a previous study by sequencing analysis. In heteroplasmic samples, minor allele proportions down to 9% were unambiguously detected and quantified. The variation in allele proportion estimates was essentially the same within and between different rtPCR runs, and the differences between total copy number estimates found for rerun samples were comparable to those found with non-allele-discriminatory quantitative rtPCR assays.

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Section: Temporal Reproducibilitysupporting

confidence: 86%

Characterization of mtDNA SNP typing and mixture ratio assessment with simultaneous real-time PCR quantification of both allelic states

Niederstätter

Coble²,

Grubwieser

et al. 2005

Int J Legal Med

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“…If n t < T , then allele drop-out will occur because the signal is insufficient to be detected by the photomultiplier. Generally, when levels of DNA are <0.05 ng/μl, then estimates of quantity tend to be unreliable (26). However, newer methods based on real-time TaqMan assays (e.g.…”

Section: Resultsmentioning

confidence: 99%

A graphical simulation model of the entire DNA process associated with the analysis of short tandem repeat loci

Gill

Curran²,

Elliot³

2005

Nucleic Acids Research

116

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The use of expert systems to interpret short tandem repeat DNA profiles in forensic, medical and ancient DNA applications is becoming increasingly prevalent as high-throughput analytical systems generate large amounts of data that are time-consuming to process. With special reference to low copy number (LCN) applications, we use a graphical model to simulate stochastic variation associated with the entire DNA process starting with extraction of sample, followed by the processing associated with the preparation of a PCR reaction mixture and PCR itself. Each part of the process is modelled with input efficiency parameters. Then, the key output parameters that define the characteristics of a DNA profile are derived, namely heterozygote balance (Hb) and the probability of allelic drop-out p(D). The model can be used to estimate the unknown efficiency parameters, such as πextraction. ‘What-if’ scenarios can be used to improve and optimize the entire process, e.g. by increasing the aliquot forwarded to PCR, the improvement expected to a given DNA profile can be reliably predicted. We demonstrate that Hb and drop-out are mainly a function of stochastic effect of pre-PCR molecular selection. Whole genome amplification is unlikely to give any benefit over conventional PCR for LCN.

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“…Periodical renewal of the buffered fixative solution in samples stored for months or years reduces the deleterious effects of old formalin. Several studies have recently appeared with methods aimed to obtain the fixation of tissues with less damage to nucleic acids and to optimize DNA extraction and evaluation (Panaro et al 2000;Coura et al 2005;Kline et al 2005;Rivero et al 2006;Stanta et al 2006). DNA preservation in paraffin blocks is variable but probably dependent on the type of fixative solution and on the length of time of tissue storage in the fixative before paraffin embedding.…”

Section: Dna Preservationmentioning

confidence: 99%

Brain banks: benefits, limitations and cautions concerning the use of post-mortem brain tissue for molecular studies

Ferrer

Martínez

Boluda

et al. 2008

Cell Tissue Banking

146

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Brain banks are facilities providing an interface between generous donation of nervous tissues and research laboratories devoted to increase our understanding of the diseases of the nervous system, discover new diagnostic targets, and develop new strategies. Considering this crucial role, it is important to learn about the suitabilities, limitations and proper handling of individual brain samples for particular studies. Several factors may interfere with preservation of DNA, RNA, proteins and lipids, and, therefore, special care must be taken first to detect sub-optimally preserved tissues and second to provide adequate material for each specific purpose. Basic aspects related with DNA, RNA and protein preservation include agonal state, post-mortem delay, temperature of storage and procedures of tissue preservation. Examination of DNA and RNA preservation is best done by using bioanalyzer technologies instead of less sensitive methods such as agarose gels. Adequate RNA preservation is mandatory in RNA microarray studies and adequate controls are necessary for proper PCR validation. Like for RNA, the preservation of proteins is not homogeneous since some molecules are more vulnerable than others. This aspect is crucial in the study of proteins including expression levels and possible post-translational modifications. Similarly, the reliability of functional and enzymatic studies in human post-mortem brain largely depends on protein preservation. Much less is known about other aspects, such as the effects of putative deleterious factors on epigenetic events such as methylation of CpGs in gene promoters, nucleosome preservation, histone modifications, and conservation of microRNA species. Most brains are appropriate for morphological approaches but not all brains are useful for certain biochemical and molecular studies.

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Results from the NIST 2004 DNA Quantitation Study

Cited by 57 publications

References 28 publications

Characterization of mtDNA SNP typing and mixture ratio assessment with simultaneous real-time PCR quantification of both allelic states

Characterization of mtDNA SNP typing and mixture ratio assessment with simultaneous real-time PCR quantification of both allelic states

A graphical simulation model of the entire DNA process associated with the analysis of short tandem repeat loci

Brain banks: benefits, limitations and cautions concerning the use of post-mortem brain tissue for molecular studies

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