Development and validation of InnoQuant™, a sensitive human DNA quantitation and degradation assessment method for forensic samples using high copy number mobile elements Alu and SVA

Pineda, G.; Montgomery, Anne; Thompson, Robyn; Indest, Brooke; Carroll, Marion L.; Sinha, Sudhir K.

doi:10.1016/j.fsigen.2014.08.007

Cited by 31 publications

(15 citation statements)

References 20 publications

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“…qPCR is routinely used to measure the concentration of total human and total human male DNA [2][3][4]. The inclusion of an additional quantification target to assess DNA degradation in forensic samples has also been used [5][6][7][8][9].…”

Section: Introductionmentioning

confidence: 99%

“…The primers and hydrolysis probe (labeled with TMR/BHQ 1 -2) for the internal PCR control (IPC) are used to amplify and detect a novel DNA template. The IPC's quantification cycle (Cq) provides information about the presence of PCR inhibitors in a DNA sample [2][3][4][5][6][7][8][9]14]. To increase susceptibility to PCR inhibitors relative to the other targets in the PowerQuant 1 System, including the 294 bp degradation amplicon, the IPC is the longest amplicon (435 bp).…”

Section: Introductionmentioning

confidence: 99%

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Human DNA quantification and sample quality assessment: Developmental validation of the PowerQuant ¨r) system

Ewing

Thompson

McLaren

et al. 2016

Forensic Science International: Genetics

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Quantification of the total amount of human DNA isolated from a forensic evidence item is crucial for DNA normalization prior to short tandem repeat (STR) DNA analysis and a federal quality assurance standard requirement. Previous commercial quantification methods determine the total human DNA and total human male DNA concentrations, but provide limited information about the condition of the DNA sample. The PowerQuant(®) System includes targets for quantification of total human and total human male DNA as well as targets for evaluating whether the human DNA is degraded and/or PCR inhibitors are present in the sample. A developmental validation of the PowerQuant(®) System was completed, following SWGDAM Validation Guidelines, to evaluate the assay's specificity, sensitivity, precision and accuracy, as well as the ability to detect degraded DNA or PCR inhibitors. In addition to the total human DNA and total human male DNA concentrations in a sample, data from the degradation target and internal PCR control (IPC) provide a forensic DNA analyst meaningful information about the quality of the isolated human DNA and the presence of PCR inhibitors in the sample that can be used to determine the most effective workflow and assist downstream interpretation.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Human DNA quantification and sample quality assessment: Developmental validation of the PowerQuant ¨r) system

Ewing

Thompson

McLaren

et al. 2016

Forensic Science International: Genetics

View full text Add to dashboard Cite

show abstract

“…The degree of DNA degradation or fragmentation can be displayed as a degradation index (DI), where DNA quantity of the short target (80 bp) is divided by DNA quantity of the long target (207 or 214 bp) . Although DI can be accurately determined using a commercial DNA quantitation kit, the ratio of the mean peak height of the short amplicons to the mean peak height of the long amplicons can be utilized to roughly determine DI .…”

Section: Resultsmentioning

confidence: 99%

Differential pre‐amplification of STR loci for fragmented forensic DNA profiling

et al. 2016

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DNA profiling of short tandem repeats (STR) has been successfully used for the identification of individuals in forensic samples, accidents and natural disasters. However, STR profiling of DNA isolated from old crime scenes and damaged biological samples is difficult due to DNA degradation and fragmentation. Here, we show that pre-amplification of STR loci using biotinylated primers for the STR loci is an efficient strategy to obtain STR profiling results from fragmented forensic samples. Analysis of STR loci with longer amplicon sizes is generally hampered, since these relatively long loci are vulnerable to DNA fragmentation. This problem was overcome by using reduced or increased primer concentrations for loci with shorter or longer amplicon sizes, respectively, in our pre-amplification strategy. In addition, pre-amplification of STR loci into two groups of short or long amplicon size increases the efficiency of STR profiling from highly fragmented forensic DNA samples. Therefore, differential pre-amplification of STR loci is an effective way to obtain DNA profiling results from fragmented forensic samples.

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“…A similar approach to evaluating a sample for degradation was more recently developed by Sinha et al . The new qPCR kit called Innoquant™, utilizes two independent genomic targets.…”

Section: Real‐time Pcr Dna‐based Human Target‐specific Methodsmentioning

confidence: 99%

Advances in forensic DNA quantification: A review

2014

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This review focuses upon a critical step in forensic biology: detection and quantification of human DNA from biological samples. Determination of the quantity and quality of human DNA extracted from biological evidence is important for several reasons. Firstly, depending on the source and extraction method, the quality (purity and length), and quantity of the resultant DNA extract can vary greatly. This affects the downstream method as the quantity of input DNA and its relative length can determine which genotyping procedure to use-standard short-tandem repeat (STR) typing, mini-STR typing or mitochondrial DNA sequencing. Secondly, because it is important in forensic analysis to preserve as much of the evidence as possible for retesting, it is important to determine the total DNA amount available prior to utilizing any destructive analytical method. Lastly, results from initial quantitative and qualitative evaluations permit a more informed interpretation of downstream analytical results. Newer quantitative techniques involving real-time PCR can reveal the presence of degraded DNA and PCR inhibitors, that provide potential reasons for poor genotyping results and may indicate methods to use for downstream typing success. In general, the more information available, the easier it is to interpret and process the sample resulting in a higher likelihood of successful DNA typing. The history of the development of quantitative methods has involved two main goals-improving precision of the analysis and increasing the information content of the result. This review covers advances in forensic DNA quantification methods and recent developments in RNA quantification.

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Development and validation of InnoQuant™, a sensitive human DNA quantitation and degradation assessment method for forensic samples using high copy number mobile elements Alu and SVA

Cited by 31 publications

References 20 publications

Human DNA quantification and sample quality assessment: Developmental validation of the PowerQuant ¨r) system

Human DNA quantification and sample quality assessment: Developmental validation of the PowerQuant ¨r) system

Differential pre‐amplification of STR loci for fragmented forensic DNA profiling

Advances in forensic DNA quantification: A review

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