An Exploratory Time Since Deposition Analysis of Whole Blood Using Metrics of DNA Degradation and Visible Absorbance Spectroscopy

Stotesbury, Theresa; Cossette, Marie‐Laurence; Newell-Bell, Tamara; Shafer, Aaron B. A.

doi:10.1007/s00024-020-02494-0

Cited by 20 publications

(30 citation statements)

References 28 publications

(32 reference statements)

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“…The addition of biological replicate as a random effect in the model only marginally increased the model fit (Table 2), which was driven by a temperature-donor effect (Table 3). This is important because previous DNA degradation models found a large effect of biological replicate [29], but the standardized nature of RINe makes it a universally comparable metric. Blood composition varies by individual, for example, research on human blood showed that the amount of RNA in healthy subjects varied from 6.7 to 22.7 μg/ml [59], this variability could lead to significant differences in concentrations of total RNA obtained from biological replicates, as we observed during our study (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Further, for both concentration and quality metrics, we predicted that the greatest decrease would occur before the complete drying of the bloodstain, as the blood still represents a hydrated environment, permitting greater degradation of larger fragments, namely 18S and 28S rRNA, via ubiquitously present RNases [25]. If this occurs, it will provide greater resolution for early time periods than previous models that have focused on DNA [23,24], as DNA is more resistant to degradation than RNA [25].…”

Section: Introductionmentioning

confidence: 96%

“…Ex-vivo, blood undergoes a series of time-dependent physicochemical and morphological changes, such as hemoglobin oxidation and conformational changes [10,16], drying [17,18], clotting [19,20], and DNA and ribonucleic acid (RNA) degradation [3,[21][22][23][24]. RNA degradation, specifically total RNA degradation, has not been studied as extensively as individual mRNA molecules [25].…”

Section: Introductionmentioning

confidence: 99%

“…The copyright holder for this preprint (which this version posted April 28, 2021. ; https://doi.org/10.1101/2021.04.28.441847 doi: bioRxiv preprint 7 [25]. If this occurs, it will provide greater resolution for early time periods than previous models that have focused on DNA [23,24], as DNA is more resistant to degradation than RNA [25].…”

Section: Introductionmentioning

confidence: 99%

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Using total RNA quality metrics for time since deposition estimates in degrading bloodstains

Elliott

Stotesbury

Shafer

2021

Preprint

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Determining the age, or time since deposition (TSD), of bloodstains would provide forensic scientists with critical information regarding the timeline of the events of bloodshed during a crime. The physicochemical changes that occur to major biomolecules as a bloodstain dries can be used to estimate the TSD of bloodstains. For example, high-resolution automated gel electrophoresis can be used to quantify the timewise degradation of DNA present in bloodstains. Our study aims to analyze and quantify the timewise degradation trends found in total RNA from bloodstains, expanding the scope of the TSD research which has previously explored DNA and targeted mRNA molecules. Fifty bloodstains were stored in plastic microcentrifuge tubes at room temperature and tested over 10 different timepoints spanning one week. A total of eight RNA metrics were visually assessed and quantified using linear regression. RNA Integrity Number equivalent (RINe), total RNA concentration, and 28S/18S rRNA peak area ratios were retained for further analyses based on their relationship with time and limited correlations. RINe and total RNA concentration both exhibited negative trends over time, highlighting a decrease in quality and quantity. RINe was the RNA metric that demonstrated the greatest association with time (R2 = 0.696). Generalized linear mixed-effects models including donor (biological replicate) as a random effect increased the fit for all RNA metrics to varying degrees, but no significant differences were found between biological replicates for the RINe metric. Our results illustrated the presence of a significant decrease in the retained RNA metrics after 24 hours, suggesting that this method could be used to reliably differentiate day-old bloodstains from older bloodstains. Future work should focus on recreating this study in different environmental conditions, including testing on a variety of substrates.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 96%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Using total RNA quality metrics for time since deposition estimates in degrading bloodstains

Elliott

Stotesbury

Shafer

2021

Preprint

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“…Weber and Lednev [121] provided an overview of what is known about the biochemical mechanisms of blood aging, how these are used to determine the age of a bloodstain, the current techniques for bloodstain age determination, emerging spectroscopic techniques and technology, and the limitations of all these methodologies. Stotesbury et al [122] and Cossette et al [123] presented further research on using visible absorbance spectroscopy and DNA degradation to explore their use in determining TSD of blood, with the latter publication focusing on the impact of temperature. Bird et al [124] considered degradation patterns of semen-specific mRNA genes to assist with determining TSD of semen.…”

Section: Time Since Depositionmentioning

confidence: 99%

DNA Transfer in Forensic Science: Recent Progress towards Meeting Challenges

Oorschot

Meakin

Kokshoorn

et al. 2021

Genes

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Understanding the factors that may impact the transfer, persistence, prevalence and recovery of DNA (DNA-TPPR), and the availability of data to assign probabilities to DNA quantities and profile types being obtained given particular scenarios and circumstances, is paramount when performing, and giving guidance on, evaluations of DNA findings given activity level propositions (activity level evaluations). In late 2018 and early 2019, three major reviews were published on aspects of DNA-TPPR, with each advocating the need for further research and other actions to support the conduct of DNA-related activity level evaluations. Here, we look at how challenges are being met, primarily by providing a synopsis of DNA-TPPR-related articles published since the conduct of these reviews and briefly exploring some of the actions taken by industry stakeholders towards addressing identified gaps. Much has been carried out in recent years, and efforts continue, to meet the challenges to continually improve the capacity of forensic experts to provide the guidance sought by the judiciary with respect to the transfer of DNA.

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Whole-genome sequencing of artificial single-nucleotide variants induced by DNA degradation in biological crime scene traces

et al. 2022

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The aim of this study was to identify artificial single-nucleotide variants (SNVs) in degraded trace DNA samples. In a preliminary study, blood samples were stored for up to 120 days and whole-genome sequencing was performed using the Snakemake workflow dna-seq-gatk-variant-calling to identify positions that vary between the time point 0 sample and the aged samples. In a follow-up study on blood and saliva samples stored under humid and dry conditions, potential marker candidates for the estimation of the age of a blood stain (= time since deposition) were identified. Both studies show that a general decrease in the mean fragment size of the libraries over time was observed, presumably due to the formation of abasic sites during DNA degradation which are more susceptible to strand breaks by mechanical shearing of DNA. Unsurprisingly, an increase in the number of failed genotype calls (no coverage) was detected over time. Both studies indicated the presence of artificial SNVs with the majority of changes happening at guanine and cytosine positions. This confirms previous studies and can be explained by depurination through hydrolytic attacks which more likely deplete guanine while deamination leads to cytosine to thymine variants. Even complete genotype switches from homozygote 0/0 genotypes to the opposite 1/1 genotypes were observed. While positions with such drastic changes might provide suitable candidate markers for estimating short-term time since deposition (TsD), 11 markers were identified which show a slower gradual change of the relative abundance of the artificial variant in both blood and saliva samples, irrespective of storage conditions.

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An Exploratory Time Since Deposition Analysis of Whole Blood Using Metrics of DNA Degradation and Visible Absorbance Spectroscopy

Cited by 20 publications

References 28 publications

Using total RNA quality metrics for time since deposition estimates in degrading bloodstains

Using total RNA quality metrics for time since deposition estimates in degrading bloodstains

DNA Transfer in Forensic Science: Recent Progress towards Meeting Challenges

Whole-genome sequencing of artificial single-nucleotide variants induced by DNA degradation in biological crime scene traces

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