Improved Forensic DNA Analysis Through the use of Alternative DNA Polymerases and Statistical Modeling of DNA Profiles

Hedman, Jonas; Nordgaard, Anders; Rasmusson, Birgitta; Ansell, Ricky; Rådström, Peter

doi:10.2144/000113246

Cited by 55 publications

(49 citation statements)

References 30 publications

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“…The Quantifiler kit includes an internal PCR control (IPC) for estimation of PCR inhibition. However, the IPC only flagged for inhibition in 14 of the 42 crime scene stains that were clearly inhibited in standard AmpFlSTR SGM Plus analysis (results not shown), strengthening the notion that the multiplex STR analysis is more sensitive to inhibitors than the Quantifiler assay [6].…”

Section: Samplesmentioning

confidence: 81%

“…The previously described forensic DNA profile index (FI) [6,21] was used to quantify the quality differences between DNA profiles generated by the different DNA polymerase-buffer systems. In short, the FI uses principal components analysis to create one single value describing the quality of an electropherogram/DNA profile based on three measures: (i) the total peak height (TPH) of the CE electropherogram, that is, the sum of the heights of all true allelic STR peaks observed in a DNA profile given in relative fluorescence units (RFU); (ii) the mean local balance (MLB, intralocus balance), that is, the mean of ratios between the two peaks within each heterozygous STR locus in an electropherogram; and (iii) the Shannon entropy (SH, interloci balance), that is, discrepancies between the sums of the peak heights of the different STR markers in an electropherogram.…”

Section: Multiplex Str Analysismentioning

confidence: 99%

“…wherer is a stable estimate of r from a calibration set of measurements [6], z is the appropriate quantile of the normal distribution based on the number of comparisons made to reach the final conclusion (based on the family level of significance), and n 1 and n 2 are the numbers of replicate values for each DNA polymerase-buffer system. If the FIgm ratio surpasses the LSR for a certain sample, there is a significant difference in quality between the DNA profiles generated by the two DNA polymerase-buffer systems.…”

Section: Multiplex Str Analysismentioning

confidence: 99%

“…However, the success rate of diagnostic PCR analysis is lowered by the presence of PCRinhibitory substances [2], a problem especially prominent in forensic DNA analysis due to the often complex sampling environment at crime scenes [3][4][5]. Recently, we reported that 12% of volume crime saliva stains with adequate DNA amounts analyzed at our laboratory still produced negative DNA profiles due to the presence of inhibitors [6]. Failure to generate acceptable DNA profiles can have severe effects; for example, it can lead to cases being left unsolved or to wrongly accused people not being exculpated.…”

Section: Introductionmentioning

confidence: 99%

“…In several studies, AmpliTaq Gold has shown lower tolerance to PCR-inhibitory substances compared with other DNA polymerases [7][8][9][10][11]. We have previously shown that short tandem repeat (STR) DNA analysis of complex crime scene samples can be improved by replacing AmpliTaq Gold with either of the alternative DNA polymerase-buffer systems Bio-X-Act Short (Bioline, London, UK), ExTaq Hot Start (TaKaRa Bio, Shiga, Japan), or PicoMaxx High Fidelity (Stratagene, La Jolla, CA, USA) [6].…”

Section: Introductionmentioning

confidence: 99%

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Synergy between DNA polymerases increases polymerase chain reaction inhibitor tolerance in forensic DNA analysis

Hedman

Nordgaard

Dufva

et al. 2010

Analytical Biochemistry

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a b s t r a c tThe success rate of diagnostic polymerase chain reaction (PCR) analysis is lowered by inhibitory substances present in the samples. Recently, we showed that tolerance to PCR inhibitors in crime scene saliva stains can be improved by replacing the standard DNA polymerase AmpliTaq Gold with alternative DNA polymerase-buffer systems (Hedman et al., BioTechniques 47 (2009) 951-958). Here we show that blending inhibitor-resistant DNA polymerase-buffer systems further increases the success rate of PCR for various types of real crime scene samples showing inhibition. For 34 of 42 ''inhibited" crime scene stains, the DNA profile quality was significantly improved using a DNA polymerase blend of ExTaq Hot Start and PicoMaxx High Fidelity compared with AmpliTaq Gold. The significance of the results was confirmed by analysis of variance. The blend performed as well as, or better than, the alternative DNA polymerases used separately for all tested sample types. When used separately, the performance of the DNA polymerases varied depending on the nature of the sample. The superiority of the blend is discussed in terms of complementary effects and synergy between the DNA polymerase-buffer systems.

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Section: Samplesmentioning

confidence: 81%

Section: Multiplex Str Analysismentioning

confidence: 99%

Section: Multiplex Str Analysismentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Synergy between DNA polymerases increases polymerase chain reaction inhibitor tolerance in forensic DNA analysis

Hedman

Nordgaard

Dufva

et al. 2010

Analytical Biochemistry

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Selective enrichment of STRs for applications in forensic human identification

2015

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Forensic human identification (HID) is currently based on determining repeat length polymorphisms located in short tandem repeat regions in the human genome. Despite the great progress made in the area of multiplex PCR-based approaches, limitations associated with challenging forensic samples such as DNA degradation, cooccurrence of inhabited microbial DNA and PCR inhibitors significantly affect the success rate of human DNA profiling. We have developed a sequence-specific pre-PCR STR enrichment method and evaluated its efficacy using DNA samples doped with various contaminants in view of its application on compromised forensic samples. This strategy has enabled us to generate complete and reproducible DNA profiles from samples doped with fivefold excess of nonhuman DNA and three to fourfold excess of various potent PCR inhibitors than that is claimed to be tolerated by some of the widely used commercial multiplex STR kits, from as little as two nanograms of degraded human DNA. The "hybrid capture"-based STR enrichment strategy described in this study is easily adaptable and offers a sensitive, efficient, and economical approach for successful human DNA profiling from compromised and recalcitrant forensic samples that are usually encountered in mass disaster incidents and missing persons' identifications.

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Developmental validation of a novel 8‐dye fluorescent typing system with 59 Y‐STRs and 3 Y‐indels for forensic applications

Liu,

Chen

et al. 2023

Electrophoresis

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An 8‐dye fluorescence‐labeling forensic Y‐chromosomal short tandem repeats (Y‐STRs) kit, the 62‐plex Y‐STR multiplex amplification system, was developed and optimized. The system was validated by testing PCR conditions, stutter ratios (SR) and peak height ratios, sensitivity, mixture samples, precision and accuracy, species‐specificity, and inhibition studies according to the Scientific Working Group on DNA Analysis Methods guidelines. PCR‐based studies showed that the recommended PCR conditions were optimized for this kit. In the sensitivity study, a full profile was obtained from template DNA with a quantity of u125 pg. Consistent profiles were obtained from three different laboratories. The SRs in all loci were less than 15%, and nice balance and suitable average peak height were shown. No peaks were detected in the profiles of common animal species and microorganisms. In the male–male mixture studies, all loci were observed at a ratio of 1:8, and in the male–female mixture study, all alleles could be profiled at a ratio of 1:500 if the male DNA inputs were ≥0.5 ng/µL. An inhibitor study demonstrated that the kit had varying degrees of resistance to the presence of common inhibitors. Population study demonstrated the 62‐plex Y‐STR Kit improved the power of discrimination in unrelated Chinese Han males (n = 192). When haplotype diversity was 1, the probability of discrimination power of the 62‐plex Y‐STR Kit was 0.9948, which is suitable for forensic investigations. The results show that the developed 8‐dye fluorescence labeling 62 loci system is sensitive, robust, convenient, and highly informative for forensic applications.

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Improved Forensic DNA Analysis Through the use of Alternative DNA Polymerases and Statistical Modeling of DNA Profiles

Cited by 55 publications

References 30 publications

Synergy between DNA polymerases increases polymerase chain reaction inhibitor tolerance in forensic DNA analysis

Synergy between DNA polymerases increases polymerase chain reaction inhibitor tolerance in forensic DNA analysis

Selective enrichment of STRs for applications in forensic human identification

Developmental validation of a novel 8‐dye fluorescent typing system with 59 Y‐STRs and 3 Y‐indels for forensic applications

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