The Experts below are selected from a list of 1485 Experts worldwide ranked by ideXlab platform
Nunzianda Frascione - One of the best experts on this subject based on the ideXlab platform.
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accumulation of endogenous and exogenous nucleic acids in Touch DNA components on hands
Electrophoresis, 2021Co-Authors: Julia Burrill, Rachel Hotta, Barbara Daniel, Nunzianda FrascioneAbstract:Successful forensic DNA profiling from handled items is increasingly routine in casework. This "Touch DNA" is thought to contain both cellular and acellular nucleic acid sources. However, there is little clarity on the origins or characteristics of this material. The cellular component consists of anucleate, terminally differentiated corneocytes (assumed to lack DNA), and the occasional nucleated cell. The acellular DNA source is fragmentary, presumably cell breakdown products. This study examines the relative contributions each component makes to the hand-secretions (endogenous) and hand-accumulations (exogenous) by recovering rinses from the inside and outside of worn gloves. Additionally, cellular and acellular DNA was measured at timepoints up to 2 h after hand washing, both with and without interim contact. Microscopic examination confirmed cell morphology and presence of nucleic acids. Following the novel application of a hair keratinocyte lysis method and plasma-DNA fragment purification to hand rinse samples, DNA profiles were generated from both fractions. Exogenous cell-free DNA is shown to be a significant source of Touch DNA, which reaccumulates quickly, although its amplifiable nuclear alleles are limited. Endogenous DNA is mostly cellular in origin and provides more allelic information consistently over time.
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accumulation of endogenous and exogenous nucleic acids in Touch DNA components on hands
Electrophoresis, 2021Co-Authors: Julia Burrill, Rachel Hotta, Barbara Daniel, Nunzianda FrascioneAbstract:Successful forensic DNA profiling from handled items is increasingly routine in casework. This "Touch DNA" is thought to contain both cellular and acellular nucleic acid sources. However, there is little clarity on the origins or characteristics of this material. The cellular component consists of anucleate, terminally differentiated corneocytes (assumed to lack DNA) and the occasional nucleated cell. The acellular DNA source is fragmentary, presumably cell breakdown products. This study examines the relative contributions each component makes to the hand-secretions (endogenous) and hand-accumulations (exogenous) by recovering rinses from the inside and outside of worn gloves. Additionally, cellular and acellular DNA was measured at timepoints up to 2 h after hand washing, both with and without interim contact. Microscopic examination confirmed cell morphology and presence of nucleic acids. Following the novel application of a hair keratinocyte lysis method and plasma-DNA fragment purification to hand rinse samples, DNA profiles were generated from both fractions. Exogenous cell-free DNA is shown to be a significant source of Touch DNA, which reaccumulates quickly, although its amplifiable nuclear alleles are limited. Endogenous DNA is mostly cellular in origin and provides more allelic information consistently over time. This article is protected by copyright. All rights reserved.
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corneocyte lysis and fragmented DNA considerations for the cellular component of forensic Touch DNA
Forensic Science International-genetics, 2021Co-Authors: Julia Burrill, Barbara Daniel, Elli Rammenou, Fatima Alawar, Nunzianda FrascioneAbstract:DNA deposited by individuals' hands is a routine part of forensic analysis, yet little is understood about the precise cellular contents left by handling. "Dead" skin cells known as corneocytes make up the majority of the cellular material left in Touch deposits by people's hands but are known to lack nuclei, making their DNA content ambiguous. Here we measure DNA released from anucleate corneocytes following various lysis methods to determine how much DNA may be present in these cells and how best to recover it from inside the cornified envelope. We demonstrate that enhanced lysis methods using a reducing agent and longer incubation may be valuable for hand deposit samples. Corneocyte DNA can be characterized as highly degraded based on the quantification, STR profiling and fluorescence microscopy of the cells from freshly washed hands. Purification to target shorter DNA fragments is demonstrated. DNA from the washed corneocyte cells is shown to constitute the majority of recoverable DNA with these methods. We consider the use of new methods adapted to cornified cells and fragmented DNA for future research into this sample type.
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exploration of cell free DNA cfDNA recovery for Touch deposits
Forensic Science International-genetics, 2021Co-Authors: Julia Burrill, Barbara Daniel, Anju Kombara, Nunzianda FrascioneAbstract:Although Touch deposit DNA is widely used in forensic casework, its cellular and acellular contents and their biological origins are poorly understood. There is evidence that the cell-free component of DNA deposited by handling may contribute substantial genetic information; however, most research into Touch DNA recovery does not separate cellular and cell-free fractions or seek to characterize their contents. This work is an important early step in developing methods to isolate the cfDNA from biological material deposited by handling. Size-filtration as a separation technique was determined to be prone to DNA loss, even on optimized control samples of pure ladder DNA. Centrifugal separation was optimized to determine minimum speed and time required to reliably remove all cellular debris from the material collected by rinsing donor hands. To determine if the centrifugal force risked rupturing shed corneocyte cells and releasing cellular DNA into the supernatant, DNA levels were measured, and cells were visualized microscopically before and after centrifugation of hand rinses. Heated buccal cells were used as a positive control to demonstrate cell rupture would be detected with these methods. Following the determination of a suitable separation technique, an investigation into purification methods for cfDNA was conducted. DNA recovery using three kits for plasma cfDNA, one for PCR clean-up and one for genomic DNA were assessed on both ladder DNA to simulate cfDNA fragments and on collected hand deposit supernatants from both unwashed and washed hands. Purification methods designed for recovery of short DNA fragments from plasma yielded the highest recovery percentage across sample types, with BioChain cfPure performing the best. Donors' hands were shown to shed high levels of cfDNA, which were better recovered with a method for short fragments than with a traditional genomic technique often used on Touch DNA samples.
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A review of trace “Touch DNA” deposits: Variability factors and an exploration of cellular composition
Forensic science international. Genetics, 2018Co-Authors: Julia Burrill, Barbara Daniel, Nunzianda FrascioneAbstract:Abstract The use in courtrooms of forensic DNA typing results from presumably Touched or handled items is increasing as the sensitivity of detection techniques improves. Research investigating how much DNA can be recovered from handled items, whether trace DNA can be detected under certain scenarios including varying degrees of indirect transfer, and factors which may influence these results is summarized here. Fundamentally, our current understanding of the cellular content of Touch deposits and the origins of the potential trace DNA therein is extremely limited. Possible origins include anucleate corneocytes, fragmentary cells/nuclei, nucleated epithelial cells from hands, transferred nucleated cells, and cell-free DNA. Here we review the existing evidence for each possible source and consider remaining knowledge gaps regarding forensically relevant Touch depositions.
Dan O M Bonsu - One of the best experts on this subject based on the ideXlab platform.
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comparison of isohelix and rayon swabbing systems for Touch DNA recovery from metal surfaces
Forensic Science Medicine and Pathology, 2021Co-Authors: Dan O M Bonsu, Denice Higgins, Matthew Rodie, Julianne Henry, Jeremy J AustinAbstract:A previous study evaluating two swabbing systems found that DNA was best recovered from sterile metal substrates using an Isohelix™ swab wetted with isopropyl alcohol rather than a Rayon swab with water as the wetting agent. We tested the same swabbing systems on metal (aluminum, brass, and stainless steel) and plastic substrates in a regularly Touched environment to simulate the non-deliberate transfer of Touch evidence likely seen in a casework scenario, to ascertain the performance of these swabs in an uncontrolled situation. Higher amounts of Touch DNA were recovered with Isohelix™ swabs (0.5 – 3.3 ng) compared to Rayon swabs (0.13 – 1.2 ng). The Isohelix™ swabbing system was found to significantly recover more Touch DNA (p = 0.04) from the metal substrates than the Rayon swabbing system, consistent with the findings of our previous work. The results contribute to our understanding of the impact of sample collection techniques on Touch DNA recovery from problematic metal surfaces and suggest that supplemental cleaning of substrates as a precautionary step against the spread of infections may affect Touch DNA persistence and the recovery efficiency of swabs.
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forensic Touch DNA recovery from metal surfaces a review
Science & Justice, 2020Co-Authors: Dan O M Bonsu, Denice Higgins, Jeremy J AustinAbstract:Trace evidence such as Touch (also known as contact) DNA has probative value as a vital forensic investigative tool that can lead to the identification and apprehension of a criminal. While the volume of Touch DNA evidence items submitted to forensic laboratories has significantly increased, recovery and amplification of DNA from these items, especially from metal surfaces, remains challenging. Currently little is understood with regards to the underlying mechanisms of metal-DNA interactions in the context of forensic science and how this may impact on DNA recovery. An increased understanding of these mechanisms would allow optimisation of methods to improve outcomes when sampling these materials. This paper reviews the basis of DNA binding to metal substrates, the merits and limitations of current methods and future perspectives of improving recovery and amplification of Touch DNA from metal surfaces of forensic interest.
Adrian Linacre - One of the best experts on this subject based on the ideXlab platform.
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evaluation of a fluorescent dye to visualize Touch DNA on various substrates
Journal of Forensic Sciences, 2021Co-Authors: Jessica Champion, Duncan Taylor, Roland A H Van Oorschot, Piyamas Kanokwongnuwut, Adrian LinacreAbstract:A wide variety of items are submitted as evidence in a forensic investigation. Identifying the location of DNA on such items is central to maximizing DNA profiling success and thus the ability to link a person of interest to a particular item or crime. Recent publications describe a fluorescent staining method using Diamond™ Dye (DD) to visualize cellular material on the surface of non-porous items (e.g., glass, plastic). However, substrates of varying porosity and background color have not yet been examined. Varying porous substrates (i.e., paper bank note, stamp, cigarette, wooden matchstick, and fabric) and non-porous substrates (i.e., enamel tooth and plastic bank note) were examined for their suitability with the use of DD. To improve the visualization of cellular material on the porous substrates, we also explored two DD diluents and adjusting image contrast. The results suggest the optimal diluent depends on the absorbent nature of the substrate. For example, ethanol was sufficient for visualization on the non-porous substrates, whereas water was better for the porous substrates. While cellular material was detected on the paper bank note, tooth, and stamp, background fluorescence or autofluorescence and surface type of matchstick prevented clear visualization on this substrate. It was also determined that by adjusting the contrast of images for tooth, paper bank note and matchstick aided in the detection of cellular material. Overall, this study extends the use of DD for latent DNA detection to absorbent substrates, highlights the limitations associated with these substrate types, and suggests modifications to improve visualization on these challenging substrates.
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Ancestry and phenotype predictions from Touch DNA using massively parallel sequencing
International Journal of Legal Medicine, 2020Co-Authors: Jennifer M Young, D. Power, P. Kanokwongnuwut, Adrian LinacreAbstract:Direct PCR can be used to successfully generate full STR profiles from DNA present on the surface of objects. STR profiles are only of use in cases where a potential donor profile is available for comparison, and DNA is of sufficient DNA quality and quantity to generate a reliable profile. Often, no donor information is available and only trace DNA is present on items. As a result, alternative techniques are required to generate genetic data that can provide investigative leads. Massively parallel sequencing (MPS) offers the ability to detect trace levels of DNA and improve DNA analysis success from Touched items. Here, we present the first application of direct PCR coupled with MPS to generate forensic intelligence SNP data from latent DNA. The panels assessed are (1) the HIrisplex System that targets 24 SNPs to simultaneously predict hair and eye, and (2) the Precision ID Ancestry Panel that targets 165 autosomal SNPs indicative of biogeographic ancestry. For each panel, we analysed 60 Touched samples across five individuals and four substrates (glass slide, fuse, zip-lock bag and wire) using Ion AmpliSeq Library Preparation Kit on the automated Ion Chef System and Ion Torrent PGM. We examine the SNP recovery, concordance with reference samples and the genotype reproducibility from different substrates and donors. The results demonstrate the application of this approach for obtaining informative genetic from trace amounts of DNA.
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use of a spray device to locate Touch DNA on casework samples
Journal of Forensic Sciences, 2020Co-Authors: Jennifer M Young, Adrian LinacreAbstract:The use of a fluorescent dye to visualize cellular material on surfaces offers a targeted sampling approach for locating Touch DNA on casework items. However, the current application of such dye is not feasible for examination of relatively large items. As a result, development of an efficient dye application system is required to translate this approach into practice. Here, the spray pattern (area covered, intensity, and evenness) of 15 different commercial spray devices was examined visually using food coloring. From this, five devices were selected to apply Diamond Nucleic Acid Dye (DD) to three substrates (glass slide, plastic sheet, and brown packing tape) seeded with saliva and Touch DNA. The cellular material was visualized using the Dino-lite Microscope and Polilight. The inhibitory effects of DD afforded by each spray device were examined using Identifiler Plus® DNA profiling kit and a DNA input of 800 pg. The two most promising devices were further tested on a range of mock casework items seeded with Touch DNA. The results presented demonstrate the feasibility of a spray system to apply DD to large surfaces and subsequently detect cellular material at both micro and macroscale. Specifically, the data suggest that a pressurized continuous-spray system is favorable and that droplet size influences the intensity of fluorescence and surface coverage. Furthermore, this study indicates that full STR profiles can be obtained following spraying with DD solution, even with excessive application, which is essential for the widespread use of these devices in casework.
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DNA profiles generated from a range of Touched sample types
Forensic Science International-genetics, 2018Co-Authors: Belinda Martin, Duncan Taylor, Renee Blackie, Adrian LinacreAbstract:Abstract Direct PCR from Touch DNA has a range of potential applications in the field of forensic investigation for exhibit examination that, under standard extraction methods, rarely produce informative DNA profiles. Previous studies from ‘Touch DNA’ have focussed on fingermarks created under laboratory conditions. Here we report on successful STR DNA profiling from a range of Touched items. Direct PCR, with no increase in cycle number, was performed after eight different sample types, typical of those submitted for forensic investigation, were handled by volunteers for a maximum of 15 s to deposit trace amounts of their DNA. Amplifications were performed using either GlobalFiler® or Identifiler® Plus following manufacturer’s instructions. These two kits were chosen deliberately as many laboratories worldwide have adopted and validated them in their workflow, thus allowing for direct PCR to be incorporated within their practises easily. It was found that informative STR profiles were obtained from all eight substrates using both STR kits. Identifiler® Plus out-performed GlobalFiler® in terms of the percentage of alleles amplified using the direct PCR approach. Both generated informative profiles from all items and all individuals, at different rates, with Identifiler® Plus being informative in a larger percentage of samples. GlobalFiler® produced profiles with an average of 60% ± 24% (36 ± 15 alleles) alleles present while Identifiler® Plus produced profiles with an average of 96% ± 4% (31 ± 1 alleles) alleles present. A comparison was made between the direct PCR approach and subjecting Touched samples to a standard DNA extraction process, both using Identifiler®. An average of 4% of profiles were informative for samples that underwent extraction with 100% being informative from the same subset of samples amplified by direct PCR. Our findings further demonstrate the success of direct PCR to enhance the STR DNA profiles from Touch DNA. Further, Identifiler® Plus was found to generate informative profiles more often than GlobalFiler®. Direct PCR is fast, simple, and non-destructive of evidence with the ability to generate informative genetic data where standard methods are likely to fail.
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genetic profiling from challenging samples direct pcr of Touch DNA
Forensic Science International: Genetics Supplement Series, 2013Co-Authors: Jennifer E L Templeton, Renee Ottens, Viviana Paradiso, Oliva Handt, Duncan Taylor, Adrian LinacreAbstract:A B S T R A C T We report on a novel direct PCR approach for retrieving Touch DNA from handled items. Trace levels of DNA on handled items can be difficult to recover and analyse using standard methodology. The lack of a full nuclear STR DNA profile may be related to insufficient DNA being present initially, the presence of PCR inhibitors, or the loss of nucleic acid during sample collection and processing. The aim of this study was to optimise the swabbing method used to collect DNA and to improve the success rate of STR-based DNA profiling from Touched items. A comparison was carried out to evaluate the use of three common swab types, nylon, cotton and foam at retrieving and releasing template DNA for amplification by direct PCR. To test the effectiveness of our method at processing low levels of DNA, complete STR DNA profiles were obtained from fingerprints deposited onto sterile plastic slides, which were swabbed and subjected to direct PCR. Our findings indicate the potential use of direct PCR when analysing samples that contain limited DNA.
Julia Burrill - One of the best experts on this subject based on the ideXlab platform.
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accumulation of endogenous and exogenous nucleic acids in Touch DNA components on hands
Electrophoresis, 2021Co-Authors: Julia Burrill, Rachel Hotta, Barbara Daniel, Nunzianda FrascioneAbstract:Successful forensic DNA profiling from handled items is increasingly routine in casework. This "Touch DNA" is thought to contain both cellular and acellular nucleic acid sources. However, there is little clarity on the origins or characteristics of this material. The cellular component consists of anucleate, terminally differentiated corneocytes (assumed to lack DNA), and the occasional nucleated cell. The acellular DNA source is fragmentary, presumably cell breakdown products. This study examines the relative contributions each component makes to the hand-secretions (endogenous) and hand-accumulations (exogenous) by recovering rinses from the inside and outside of worn gloves. Additionally, cellular and acellular DNA was measured at timepoints up to 2 h after hand washing, both with and without interim contact. Microscopic examination confirmed cell morphology and presence of nucleic acids. Following the novel application of a hair keratinocyte lysis method and plasma-DNA fragment purification to hand rinse samples, DNA profiles were generated from both fractions. Exogenous cell-free DNA is shown to be a significant source of Touch DNA, which reaccumulates quickly, although its amplifiable nuclear alleles are limited. Endogenous DNA is mostly cellular in origin and provides more allelic information consistently over time.
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accumulation of endogenous and exogenous nucleic acids in Touch DNA components on hands
Electrophoresis, 2021Co-Authors: Julia Burrill, Rachel Hotta, Barbara Daniel, Nunzianda FrascioneAbstract:Successful forensic DNA profiling from handled items is increasingly routine in casework. This "Touch DNA" is thought to contain both cellular and acellular nucleic acid sources. However, there is little clarity on the origins or characteristics of this material. The cellular component consists of anucleate, terminally differentiated corneocytes (assumed to lack DNA) and the occasional nucleated cell. The acellular DNA source is fragmentary, presumably cell breakdown products. This study examines the relative contributions each component makes to the hand-secretions (endogenous) and hand-accumulations (exogenous) by recovering rinses from the inside and outside of worn gloves. Additionally, cellular and acellular DNA was measured at timepoints up to 2 h after hand washing, both with and without interim contact. Microscopic examination confirmed cell morphology and presence of nucleic acids. Following the novel application of a hair keratinocyte lysis method and plasma-DNA fragment purification to hand rinse samples, DNA profiles were generated from both fractions. Exogenous cell-free DNA is shown to be a significant source of Touch DNA, which reaccumulates quickly, although its amplifiable nuclear alleles are limited. Endogenous DNA is mostly cellular in origin and provides more allelic information consistently over time. This article is protected by copyright. All rights reserved.
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corneocyte lysis and fragmented DNA considerations for the cellular component of forensic Touch DNA
Forensic Science International-genetics, 2021Co-Authors: Julia Burrill, Barbara Daniel, Elli Rammenou, Fatima Alawar, Nunzianda FrascioneAbstract:DNA deposited by individuals' hands is a routine part of forensic analysis, yet little is understood about the precise cellular contents left by handling. "Dead" skin cells known as corneocytes make up the majority of the cellular material left in Touch deposits by people's hands but are known to lack nuclei, making their DNA content ambiguous. Here we measure DNA released from anucleate corneocytes following various lysis methods to determine how much DNA may be present in these cells and how best to recover it from inside the cornified envelope. We demonstrate that enhanced lysis methods using a reducing agent and longer incubation may be valuable for hand deposit samples. Corneocyte DNA can be characterized as highly degraded based on the quantification, STR profiling and fluorescence microscopy of the cells from freshly washed hands. Purification to target shorter DNA fragments is demonstrated. DNA from the washed corneocyte cells is shown to constitute the majority of recoverable DNA with these methods. We consider the use of new methods adapted to cornified cells and fragmented DNA for future research into this sample type.
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exploration of cell free DNA cfDNA recovery for Touch deposits
Forensic Science International-genetics, 2021Co-Authors: Julia Burrill, Barbara Daniel, Anju Kombara, Nunzianda FrascioneAbstract:Although Touch deposit DNA is widely used in forensic casework, its cellular and acellular contents and their biological origins are poorly understood. There is evidence that the cell-free component of DNA deposited by handling may contribute substantial genetic information; however, most research into Touch DNA recovery does not separate cellular and cell-free fractions or seek to characterize their contents. This work is an important early step in developing methods to isolate the cfDNA from biological material deposited by handling. Size-filtration as a separation technique was determined to be prone to DNA loss, even on optimized control samples of pure ladder DNA. Centrifugal separation was optimized to determine minimum speed and time required to reliably remove all cellular debris from the material collected by rinsing donor hands. To determine if the centrifugal force risked rupturing shed corneocyte cells and releasing cellular DNA into the supernatant, DNA levels were measured, and cells were visualized microscopically before and after centrifugation of hand rinses. Heated buccal cells were used as a positive control to demonstrate cell rupture would be detected with these methods. Following the determination of a suitable separation technique, an investigation into purification methods for cfDNA was conducted. DNA recovery using three kits for plasma cfDNA, one for PCR clean-up and one for genomic DNA were assessed on both ladder DNA to simulate cfDNA fragments and on collected hand deposit supernatants from both unwashed and washed hands. Purification methods designed for recovery of short DNA fragments from plasma yielded the highest recovery percentage across sample types, with BioChain cfPure performing the best. Donors' hands were shown to shed high levels of cfDNA, which were better recovered with a method for short fragments than with a traditional genomic technique often used on Touch DNA samples.
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A review of trace “Touch DNA” deposits: Variability factors and an exploration of cellular composition
Forensic science international. Genetics, 2018Co-Authors: Julia Burrill, Barbara Daniel, Nunzianda FrascioneAbstract:Abstract The use in courtrooms of forensic DNA typing results from presumably Touched or handled items is increasing as the sensitivity of detection techniques improves. Research investigating how much DNA can be recovered from handled items, whether trace DNA can be detected under certain scenarios including varying degrees of indirect transfer, and factors which may influence these results is summarized here. Fundamentally, our current understanding of the cellular content of Touch deposits and the origins of the potential trace DNA therein is extremely limited. Possible origins include anucleate corneocytes, fragmentary cells/nuclei, nucleated epithelial cells from hands, transferred nucleated cells, and cell-free DNA. Here we review the existing evidence for each possible source and consider remaining knowledge gaps regarding forensically relevant Touch depositions.
Jack Ballantyne - One of the best experts on this subject based on the ideXlab platform.
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single source DNA profile recovery from single cells isolated from skin and fabric from Touch DNA mixtures in mock physical assaults
Science & Justice, 2017Co-Authors: Katherine Farash, Erin K Hanson, Jack BallantyneAbstract:The ability to obtain DNA profiles from trace biological evidence is routinely demonstrated with so-called 'Touch DNA evidence', which is generally perceived to be the result of DNA obtained from shed skin cells transferred from a donor's hands to an object or person during direct physical contact. Current methods for the recovery of trace DNA employ swabs or adhesive tape to sample an area of interest. While of practical utility, such 'blind-swabbing' approaches will necessarily co-sample cellular material from the different individuals whose cells are present on the item, even though the individuals' cells are principally located in topographically dispersed, but distinct, locations on the item. Thus the act of swabbing itself artifactually creates some of the DNA mixtures encountered in Touch DNA samples. In some instances involving transient contact between an assailant and victim, the victim's DNA may be found in such significant excess as to preclude the detection and typing of the perpetrator's DNA. In order to circumvent the challenges with standard recovery and analysis methods for Touch DNA evidence, we reported previously the development of a 'smart analysis' single cell recovery and DNA analysis method that results in enhanced genetic analysis of Touch DNA evidence. Here we use the smart single cell analysis method to recover probative single source profiles from individual and agglomerated cells from various Touched objects and clothing items belonging to known donors. We then use the same approach for the detection of single source male donor DNA in simulated physical contact/assault mixture samples (i.e. male 'assailant' grabbing the wrist, neck or clothing from the female 'victim', or being in transient contact with bedding from the 'victim'). DNA profiles attributable to the male or female known donors were obtained from 31% and 35% of the single and agglomerated bio-particles (putative cells) tested. The known male donor 'assailant' DNA profile was identified in the cell sampling from every mixture type tested. The results of this work demonstrate the efficacy of an alternative strategy to recover single source perpetrator DNA profiles in physical contact/assault cases involving trace perpetrator/victim cellular admixtures.
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enhanced genetic analysis of single human bioparticles recovered by simplified micromanipulation from forensic Touch DNA evidence
Journal of Visualized Experiments, 2015Co-Authors: Katherine Farash, Erin K Hanson, Jack BallantyneAbstract:DNA profiles can be obtained from ‘Touch DNA’ evidence, which comprises microscopic traces of human biological material. Current methods for the recovery of trace DNA employ cotton swabs or adhesive tape to sample an area of interest. However, such a ‘blind-swabbing’ approach will co-sample cellular material from the different individuals, even if the individuals’ cells are located in geographically distinct locations on the item. Thus, some of the DNA mixtures encountered in Touch DNA samples are artificially created by the swabbing itself. In some instances, a victim’s DNA may be found in significant excess thus masking any potential perpetrator’s DNA. In order to circumvent the challenges with standard recovery and analysis methods, we have developed a lower cost, ‘smart analysis’ method that results in enhanced genetic analysis of Touch DNA evidence. We describe an optimized and efficient micromanipulation recovery strategy for the collection of bio-particles present in Touch DNA samples, as well as an enhanced amplification strategy involving a one-step 5 µl microvolume lysis/STR amplification to permit the recovery of STR profiles from the bio-particle donor(s). The use of individual or few (i.e., “clumps”) bioparticles results in the ability to obtain single source profiles. These procedures represent alternative enhanced techniques for the isolation and analysis of single bioparticles from forensic Touch DNA evidence. While not necessary in every forensic investigation, the method could be highly beneficial for the recovery of a single source perpetrator DNA profile in cases involving physical assault (e.g., strangulation) that may not be possible using standard analysis techniques. Additionally, the strategies developed here offer an opportunity to obtain genetic information at the single cell level from a variety of other non-forensic trace biological material.
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getting blood from a stone ultrasensitive forensic DNA profiling of microscopic bio particles recovered from Touch DNA evidence
Methods of Molecular Biology, 2013Co-Authors: Erin K Hanson, Jack BallantyneAbstract:In forensic casework analysis it is sometimes necessary to obtain genetic profiles from increasingly smaller amounts of biological material left behind by persons involved in criminal offenses. The ability to obtain profiles from trace biological evidence is routinely demonstrated with the so-called Touch DNA evidence (generally perceived to be the result of DNA obtained from shed skin cells transferred from donor to an object or a person during physical contact). The current method of recovery of trace DNA employs cotton swabs or adhesive tape to sample an area of interest. While of practical utility, such a "blind-swabbing" approach will necessarily co-sample cellular material from the different individuals whose cells are present on the item, even if the individuals' cells are located in geographically distinct locations on the item. Thus some of the DNA mixtures encountered in such Touch DNA samples are artificially created by the swabbing itself. Therefore, a specialized approach for the isolation of single or few cells from "Touch DNA evidence" is necessary in order to improve the analysis and interpretation of profiles recovered from these samples. Here, we describe an optimized and efficient removal strategy for the collection of cellular microparticles present in "Touch DNA" samples, as well as enhanced amplification strategies to permit the recovery of short tandem repeat profiles of the donor(s) of the recovered microparticles.
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specific and sensitive mrna biomarkers for the identification of skin in Touch DNA evidence
Forensic Science International-genetics, 2012Co-Authors: Erin K Hanson, Cordula Haas, R Jucker, Jack BallantyneAbstract:In forensic casework analysis it is often necessary to attempt to obtain DNA profiles from microscopic amounts of biological material left behind by perpetrators of crime. The ability to obtain profiles from trace biological evidence is routinely demonstrated with so-called 'Touch DNA' evidence, which is generally perceived to be the result of DNA obtained from shed skin cells transferred from donor to an object or person during physical contact. Although a genetic profile from trace biological evidence is routinely obtained, the tissue source of the profile is rarely known. This merely perpetuates the 'mystery' of the nature of 'Touch DNA' evidence allowing the significance or meaningfulness of genetic profiles obtained from these samples to be challenged. Numerous reports state that the tissue source of origin of 'Touch DNA' evidence cannot be determined due to the small amount of biological material present, while others conclude that the DNA profiles are obtained from shed skin cells (as opposed to, say, buccal epithelial cells present in saliva traces) without any scientific basis for this assertion. Proper identification of the biological material present might be crucial to the investigation and prosecution of a criminal offense and a misrepresentation of the nature of the evidence can have undue influence on the perception of the circumstance of the crime. Thus far, research has failed to provide forensic scientists with feasible, definitive methods to identify the tissue origin of 'Touch DNA'. In the present work, we sought to identify novel highly specific and sensitive messenger RNA (mRNA) biomarkers for the identification of skin. Gene candidates were identified using both literature searches and whole transcriptome deep sequencing (RNA-Seq). Utilizing this dual approach, we identified and evaluated over 100 gene candidates. Five mRNA markers were identified that demonstrated a high degree of specificity for skin. Using these markers, we have been able to successfully detect and identify skin using as little as 5-25 pg of input total RNA from skin and, significantly, in swabs of human skin and various Touched objects. One of the markers, LCE1C, is particularly highly sensitive and was detected in the majority of skin samples tested including Touched objects. We have been successful in incorporating the five skin biomarkers into two multiplex systems. Although further work is needed to optimize the assay for routine casework, the initial studies demonstrate that a molecular-based characterization of the biological material recovered from Touch samples is possible.