ORCID Profile
0000-0001-5890-5548
Current Organisation
University of Sussex
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Research Topics
In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Top 5 Research Topics
ANZSRC Field of Research (FoR)
Forensic Biology | Other Biological Sciences | Forensic Chemistry | Evolutionary Impacts of Climate Change | Genomics | Access to Justice | Evolutionary Biology | Archaeological Science |
ANZSRC Socio-Economic Objective (SEO)
Criminal Justice | Ecosystem Adaptation to Climate Change | Understanding Australia's Past | National Security | Law Enforcement | Flora, Fauna and Biodiversity at Regional or Larger Scales | Trade and Environment
Related Links
Publications
Cytochrome b or cytochrome c oxidase subunit I for mammalian species identification-An answer to the debate
Publisher: Elsevier BV
Date: 12-2009
The use of mitochondrial DNA genes to identify closely related avian species
Publisher: Elsevier BV
Date: 12-2009
A method to identify a large number of mammalian species in the UK from trace samples and mixtures without the use of sequencing
Publisher: Elsevier BV
Date: 08-2008
Quantification of trace amounts of human and non-human mitochondrial DNA (mtDNA) using SYBR Green and real time PCR
Publisher: Elsevier BV
Date: 08-2008
A rapid screening method using DNA binding dyes to determine whether hair follicles have sufficient DNA for successful profiling
Publisher: Elsevier BV
Date: 05-2016
DOI: 10.1016/J.FORSCIINT.2016.03.026
Abstract: We report a simple screening method to assess the viability of successful DNA profiling from single hair follicles. A total of 48 hair s les (shed and plucked) were collected from male and female donors and the root tips (0.5cm) were stained using one of three DNA binding dyes (EvaGreen™, Diamond™ Nucleic Acid Dye and RedSafe™) at 20× concentration. The hairs were subsequently viewed under a Nikon Optiphot fluorescent microscope to count the approximate number of nuclei in one plane of view. The hairs were then processed using either (1) a DNA extraction kit (QIAmp(®) Mini Kit) and then lified using the AmpFLSTR(®) NGM™ kit, which lifies 15 short tandem repeat (STR) loci plus the gender marker amelogenin, or (2) by direct PCR lification using the same DNA profiling kit. Diamond™ dye had the lowest background signal and plucked hairs treated with this dye produced full DNA profiles when lified directly and was chosen to screen a further 150 mixed hair s les. These hairs were separated into one of five categories (1, >100 nuclei 1.5, 50-99 nuclei 2, 1-49 nuclei 2.5, no nuclei but high fluorescent signal 3, no nuclei and very low fluorescent signal) from which 60 of the hairs were chosen to undergo direct lification using the NGM™ kit. It was found that there was a direct correlation to the category designation and the ability to obtain a DNA profile up-loadable to the Australian DNA Database. Approximately 91% of category 1 hairs resulted in either a full or high partial (12-29 alleles) profile by direct PCR whereas about 78% of category 3 hairs exhibited no lification. The results show that this method can be used to predict successful STR lification from single hair follicles. It is a rapid, sensitive, cheap, non-destructive and easy to perform methodology applicable for screening multiple hairs in order to aid forensic investigators in predicting hairs that will yield DNA results.
Identifying endangered species from degraded mixtures at low levels
Publisher: Elsevier BV
Date: 12-2009
The development of a tool to predict temperature-exposure of incinerated teeth using colourimetric and hydroxyapatite crystal size data
Publisher: Springer Science and Business Media LLC
Date: 02-03-2021
Effect of nucleic acid binding dyes on DNA extraction, amplification, and STR typing
Publisher: Wiley
Date: 10-2015
Abstract: We report on the effects of six dyes used in the detection of DNA on the process of DNA extraction, lification, and detection of STR loci. While dyes can be used to detect the presence of DNA, their use is restricted if they adversely affect subsequent DNA typing processes. Diamond™ Nucleic Acid Dye, GelGreen™, GelRed™, RedSafe™, SYBR(®) Green I, and EvaGreen™ were evaluated in this study. The percentage of dye removed during the extraction process was determined to be: 70.3% for SYBR(®) Green I 99.6% for RedSafe™ 99.4% for EvaGreen™ 52.7% for Diamond™ Dye 50.6% for GelRed™, and could not be determined for GelGreen™. It was then assumed that the amount of dye in the fluorescent quantification assay had no effect on the DNA signal. The presence of all six dyes was then reviewed for their effect on DNA extraction. The t-test showed no significant difference between the dyes and the control. These extracts were then STR profiled and all dyes and control produced full DNA profiles. STR loci in the presence of GelGreen(TM) at 1X concentration showed increased lification products in comparison to the control s les. Full STR profiles were detected in the presence of EvaGreen™ (1X), although with reduced lification products. RedSafe™ (1X), Diamond™ Dye (1X), and SYBR(®) Green I (1X) all exhibited varying degrees of locus drop-out with GelRed™ generating no loci at all. We provide recommendations for the best dye to visualize the presence of DNA profile as a biological stain and its subsequent lification and detection.
Successful direct amplification of nuclear markers from single dog hairs using DogFiler multiplex
Publisher: Wiley
Date: 14-07-2015
Abstract: We report on successful lification of canine STR DNA profiles from single dog hairs. Dog hairs are commonly found on clothing or items of interest in forensic casework and may be crucial associative evidence if linked to an in idual dog. We used direct lification from these hairs to increase the DNA yield of the s le, as well as greatly reducing analysis time. Hairs from different somatic regions were used from several different dog breeds to lify a selection of eight loci from the validated DogFiler multiplex. Naturally shed canine hairs were processed, with a mix of coarse topcoat (guard) hairs and thinner soft undercoat hairs. Multiple sections of single hairs were lified in 5 mm segments to determine the viability of DNA recovery from the shaft of the hair. Single guard hairs were cut into 5 mm sections and added directly into a PCR tube. Undercoat hairs, which are very fine, were lified together in a single tube (approximately ten small hairs). Coarse hairs were found to be the most successful in producing full DNA profiles at all eight loci, matching the corresponding reference profile for that dog.
‘ForCyt’ DNA database of wildlife species
Publisher: Elsevier BV
Date: 12-2017
DNA typing in wildlife crime: recent developments in species identification
Publisher: Springer Science and Business Media LLC
Date: 05-06-2010
DOI: 10.1007/S12024-010-9168-7
Abstract: Species identification has become a tool in the investigation of acts of alleged wildlife crimes. This review details the steps required in DNA testing in wildlife crime investigations and highlights recent developments where not only can in idual species be identified within a mixture of species but multiple species can be identified simultaneously. 'What species is this?' is a question asked frequently in wildlife crime investigations. Depending on the material being examined, DNA analysis may offer the best opportunity to answer this question. Species testing requires the comparison of the DNA type from the unknown s le to DNA types on a database. The areas of DNA tested are on the mitochondria and include predominantly the cytochrome b gene and the cytochrome oxidase I gene. Standard analysis requires the sequencing of part of one of these genes and comparing the sequence to that held on a repository of DNA sequences such as the GenBank database. Much of the DNA sequence of either of these two genes is conserved with only parts being variable. A recent development is to target areas of those sequences that are specific to a species this can increase the sensitivity of the test with no loss of specificity. The benefit of targeting species specific sequences is that within a mixture of two of more species, the in idual species within the mixture can be identified. This identification would not be possible using standard sequencing. These new developments can lead to a greater number of s les being tested in alleged wildlife crimes.
DNA profiles from clothing fibers using direct PCR
Publisher: Springer Science and Business Media LLC
Date: 16-07-2016
DOI: 10.1007/S12024-016-9784-Y
Abstract: We report on the successful use of direct PCR lification of single fibers from items of worn clothing. Items of clothing were worn throughout the course of a day, with the in idual commencing regular activities. Single fibers were taken from the cuff of the clothing at regular intervals and lified directly. The same areas were subjected to tape-lifting, and also lified directly for comparison. The NGM™ kit that lifies 15 STR loci plus amelogenin was used. A total of 35 single fiber s les were processed and analyzed from five items of clothing, with 81 % of s les returning a profile of 14 alleles or more. All tape-lift s les lified directly produced DNA profiles of 15 alleles or more. The aim was to develop a simple, operational method that could be used routinely in forensic science casework and that has the potential to generate more complete profiles, which would not be detected using standard extraction methods on this type of s le. For ease of implementation, the process also adheres to standard methods with no increase in the cycle number.
A new assay for identifying endangered species in Traditional East Asian Medicine
Publisher: Elsevier BV
Date: 12-2011
Identification multiplex assay of 19 terrestrial mammal species present in New Zealand
Publisher: Wiley
Date: 13-11-2013
Abstract: An identification assay has been developed that allows accurate detection of 19 of the most common terrestrial mammals present in New Zealand (cow, red deer, goat, dog, horse, hedgehog, cat, tammar wallaby, mouse, weasel, ferret, stoat, sheep, rabbit, Pacific rat, Norway rat, ship rat, pig, and brushtail possum). This technique utilizes species-specific primers that, combined in a multiplex PCR, target small fragments of the mitochondrial cytochrome b gene. Each species, except hedgehog, produces two distinctive species-specific fragments, making the assay self-confirmatory and enabling the identification of multiple species simultaneously in DNA mixtures. The multiplex assay detects as little as 100 copies of mitochondrial DNA, which makes it a very reliable tool for degraded and trace s les. Reliability, accuracy, reproducibility, and sensitivity tests to validate the technique were performed. The technique featured here enabled a prompt response in a predation specific event, but can also be useful for wildlife management and conservation, pest incursions detection, forensic, and industrial purposes in a very simple and cost-effective manner.
Direct identification of forensic body fluids using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry
Publisher: Elsevier BV
Date: 03-2016
Reconstructing mammalian phylogenies: a detailed comparison of the cytochrome B and cytochrome oxidase subunit I mitochondrial genes.
Publisher: Public Library of Science (PLoS)
Date: 30-11-2010
Duration of in situ fluorescent signals within hairs follicles
Publisher: Elsevier BV
Date: 12-2015
Erratum to: The complete mitochondrial genome analysis of the tiger (Panthera tigris)
Publisher: Springer Science and Business Media LLC
Date: 08-2012
Detection of latent DNA
Publisher: Elsevier BV
Date: 11-2018
DOI: 10.1016/J.FSIGEN.2018.08.004
Abstract: Touch DNA is one of the most common s le types submitted for DNA profiling. There is currently no process to visualise the presence of such DNA deposited when a person makes direct contact with items of forensic relevance. This report demonstrates the effective use of Diamond Dye to bind to DNA and allow visualisation of deposited cellular material using a mini-fluorescence microscope. Volunteers made contact with a range of items typical of those submitted as part of a forensic investigation. Contact was for less than 5 s and occurred either 15 min after hands were washed to remove any traces of DNA, and therefore under controlled conditions, or at an undefined time post handwashing to mimic real-world scenarios. Diamond Dye bound to cellular material on all the items used and in all cases it was clear where the volunteers had made this brief contact. It was also clear where no contact had been made. DNA profiling was performed on a sub-set of s les to confirm that the cellular material viewed under the microscope was human in origin and deposited by the person contacting the item this was the result obtained in every s le tested. Diamond dye is relatively inexpensive, simple to apply, binds to the DNA in 3 s or less, has no mutagenic effects at the concentrations used, does not affect subsequent DNA profiling, and does not bind effectively to bacterial DNA. In combination with a mini-fluorescence microscope, this proof-of-concept study shows that otherwise invisible DNA deposited by touch can be visualised. The position and amount of cellular material deposited during even brief contact can be recorded allowing targeted s ling in any further DNA typing of forensically-significant items.
Species identification of human and deer from mixed biological material
Publisher: Elsevier BV
Date: 07-2007
Protected DNA strand displacement for enhanced single nucleotide discrimination in double-stranded DNA
Publisher: Springer Science and Business Media LLC
Date: 04-03-2015
DOI: 10.1038/SREP08721
Where does this tiger come from?-A robust molecular technique for simultaneous identification of endangered species and subspecies
Publisher: Elsevier BV
Date: 12-2011
A multiplex assay to identify 18 European mammal species from mixtures using the mitochondrial cytochromeb gene
Publisher: Wiley
Date: 2008
Abstract: A novel species-specific multiplex to identify 18 common European mammalian species (badger, cat, cow, dog, donkey, fox, goat, guinea pig, harvest mouse, hedgehog, horse, house mouse, human, pig, rabbit, rat, red deer and sheep), many of which are often associated with forensic investigations, has been developed. The assay is based on the mitochondrial cytochrome b gene, which is commonly used in species identification and phylogeny studies. Areas of homology and variation were identified and were used to create universal and species-specific primers. The species-specific primers were designed such that they will only react with the species for which they were designed. Two primer sets were designed for each species making the test self-confirmatory. All primer sets produced the expected results. The multiplex was balanced at template concentration of 40 000 copies (approximately 1.36 pg). Validation was accomplished by analysing the same s le ten times to determine run variation and several s les for each species to determine between-s le variation. Twenty-eight additional mammalian species were reacted with the multiplex. The multiplex provides, for the first time, a definitive method for identification of species in a forensic context.
Optimising direct PCR from anagen hair samples
Publisher: Elsevier BV
Date: 2013
Current and future directions of DNA in wildlife forensic science
Publisher: Elsevier BV
Date: 05-2014
DOI: 10.1016/J.FSIGEN.2013.12.007
Abstract: Wildlife forensic science may not have attained the profile of human identification, yet the scale of criminal activity related to wildlife is extensive by any measure. Service delivery in the arena of wildlife forensic science is often ad hoc, unco-ordinated and unregulated, yet many of those currently dedicated to wildlife conservation and the protection of endangered species are striving to ensure that the highest standards are met. The genetic markers and software used to evaluate data in wildlife forensic science are more varied than those in human forensic identification and are rarely standardised between species. The time and resources required to characterise and validate each genetic maker is considerable and in some cases prohibitive. Further, issues are regularly encountered in the construction of allelic databases and allelic ladders essential in human identification studies, but also applicable to wildlife criminal investigations. Accreditation and certification are essential in human identification and are currently being strived for in the forensic wildlife community. Ex les are provided as to how best practice can be demonstrated in all areas of wildlife crime analysis and ensure that this field of forensic science gains and maintains the respect it deserves. This review is aimed at those conducting human identification to illustrate how research concepts in wildlife forensic science can be used in the criminal justice system, as well as describing the real importance of this type of forensic analysis.
On the trial of tigers–tracking tiger in Traditional East Asian Medicine
Publisher: Elsevier BV
Date: 08-2008
Toehold-Mediated Nonenzymatic DNA Strand Displacement As a Platform for DNA Genotyping
Publisher: American Chemical Society (ACS)
Date: 03-04-2013
DOI: 10.1021/JA310991R
Abstract: Toehold-mediated DNA strand displacement provides unique advantages in the construction and manipulation of multidimensional DNA nanostructures as well as nucleic acid sequence analysis. We demonstrate a step change in the use of toehold-mediated DNA strand displacement reactions, where a double-stranded DNA duplex, containing a single-stranded toehold domain, enzymatically generated and then treated as a molecular target for analysis. The approach was successfully implemented for human DNA genotyping, such as gender identification where the amelogenin gene was used as a model target system, and detecting single nucleotide polymorphisms of human mitochondrial DNA. Kinetics of the strand displacement was monitored by the quenched Förster resonance energy transfer effect.
The complete mitochondrial genome analysis of the tiger (Panthera tigris)
Publisher: Springer Science and Business Media LLC
Date: 30-12-2011
DOI: 10.1007/S11033-011-1384-Z
Abstract: The complete mitochondrial genomes of five tiger s les from three subspecies (P. t. sumatrae, P. t. altica, and P. t. tigris) were successfully obtained by using 26 specifically designed Panthera-specific primer sets. The genome organization and gene arrangement of the five tiger s les were similar to each other however polymorphic tandem repeat sequences were observed in the control region (CR). This led to a difference in the genome lengths obtained from these five s les with an average size of 16,994 bp for the five tiger mitochondrial genomes. The nucleotide base composition was on average as follows: A, 31.8% T, 27.0% C, 26.6% G, 14.6% and exhibited compositional asymmetry. Most of tiger mitochondrial genome characteristics are similar to those of other common vertebrate species however, some distinctive features were observed in the CR. First, the repetitive sequence 2 (RS 2) contained two repeat units of 80 bp and the first 15 bp of what would be the third repeat motif. The repetitive sequence 3 (RS 3) contained 47-50 repeat motifs of a shorter 8 bp (ACGTAYAC)(n). Second, length heteroplasmy polycystosine (poly-C) stretches was observed at the end of the HV I locus in all tiger s les.
Assigning confidence to sequence comparisons for species identification: A detailed comparison of the cytochrome b and cytochrome oxidase subunit I mitochondrial genes
Publisher: Elsevier BV
Date: 12-2011
Properties of nucleic acid staining dyes used in gel electrophoresis
Publisher: Wiley
Date: 23-02-2015
Abstract: Nucleic acid staining dyes are used for detecting nucleic acids in electrophoresis gels. Historically, the most common dye used for gel staining is ethidium bromide, however due to its toxicity and mutagenicity other dyes that are safer to the user and the environment are preferred. This Short Communication details the properties of dyes now available and their sensitivity for detection of DNA and their ability to permeate the cell membrane. It was found that GelRed™ was the most sensitive and safest dye to use with UV light excitation, and both GelGreen™ and Diamond™ Nucleic Acid Dye were sensitive and the safer dyes using blue light excitation.
Species Identification Using DNA Loci
Publisher: CRC Press
Date: 12-03-2009
Successful direct amplification of nuclear markers from a single hair follicle
Publisher: Springer Science and Business Media LLC
Date: 30-12-2012
DOI: 10.1007/S12024-012-9402-6
Abstract: We report on successful lification of DNA profiles from a single hair. Direct lification was used on the root tip of both anagen and telogen hairs using a kit to lify 15 STR loci. All 30 anagen hairs tested from five different people gave full DNA profiles after 29 cycles with no allelic drop-in or heterozygous imbalance. Six of the 30 telogen hairs tested resulted in a full DNA profile, and a further four telogen hair s les tested produced a DNA profile of five or more complete loci that could be up-loaded to the National DNA Database (Australia). A full DNA profile was also obtained from the shaft of an anagen hair. Current practice for many laboratories is that a single hair may not be subjected to DNA testing as there is little chance of success, hence this 100 % success rate from anagen hairs is a significant advancement. A full DNA profile was obtained from a 5 year-old single hair illustrating the success when using direct PCR rather than attempting an extraction prior to the lification step. The process described deliberately uses current DNA profiling methods with no increase in cycle number, such that the methodology can be incorporated readily into operational practice. For the first time in the field of human identification, single hairs can be analyzed with confidence that a meaningful DNA profile will be generated and the data accepted by the criminal justice system.
Profiling pythons to combat common illegal wildlife activities
Publisher: Elsevier BV
Date: 2013
Application of direct PCR in forensic casework
Publisher: Elsevier BV
Date: 2013
Optimization of Diamond Nucleic Acid Dye for quantitative PCR
Publisher: Future Science Ltd
Date: 10-2016
DOI: 10.2144/000114458
Abstract: Here, we evaluate Diamond Nucleic Acid Dye (DD) for use in quantitative PCR (qPCR) applications. Although DD is a commercially available stain for detection of DNA separated by gel electrophoresis, its use as a detection dye in qPCR has yet to be described. To determine if DD can be used in qPCR, we investigated its inhibitory effects on qPCR at concentrations ranging 0.1–2.5×. Serial dilution of DNA was used to determine the efficiency, sensitivity, and linearity of DD-generated qPCR data in comparison to other commonly used fluorescent dyes such as SYBR Green (SG), EvaGreen (EG), and BRYT Green (BG). DD was found to be comparable with other dyes for qPCR applications, with an R2 value .9 and an efficiency of 0.83. Mitochondrial DNA (mtDNA) target signals were successfully produced by DD over a DNA dilution range of ∼28 ng— 0.28 pg, demonstrating comparable sensitivity to the other dyes investigated. C q values obtained using DD were lower than those using EG by almost 7 cycles. We conclude that Diamond Nucleic Acid Dye is a cheaper, less toxic alternative for qPCR applications.
The development and validation of a single SNaPshot multiplex for tiger species and subspecies identification - Implications for forensic purposes
Publisher: Elsevier BV
Date: 03-2012
Sequence selective capture, release and analysis of DNA using a magnetic microbead-assisted toehold-mediated DNA strand displacement reaction
Publisher: Royal Society of Chemistry (RSC)
Date: 2014
DOI: 10.1039/C4AN00694A
Abstract: Oligonucleotide modified magnetic beads for the selective capture and release of forensically relevant genes for human identification.
Diagnostic models to predict nuclear DNA and mitochondrial DNA recovery from incinerated teeth
Publisher: Springer Science and Business Media LLC
Date: 12-06-2023
Amelogenin locus typing using toehold-assisted fluorescent DNA melting analysis
Publisher: Elsevier BV
Date: 2013
Typing DNA profiles from previously enhanced fingerprints using direct PCR
Publisher: Elsevier BV
Date: 07-2017
DOI: 10.1016/J.FSIGEN.2017.05.006
Abstract: Fingermarks are a source of human identification both through the ridge patterns and DNA profiling. Typing nuclear STR DNA markers from previously enhanced fingermarks provides an alternative method of utilising the limited fingermark deposit that can be left behind during a criminal act. Dusting with fingerprint powders is a standard method used in classical fingermark enhancement and can affect DNA data. The ability to generate informative DNA profiles from powdered fingerprints using direct PCR swabs was investigated. Direct PCR was used as the opportunity to generate usable DNA profiles after performing any of the standard DNA extraction processes is minimal. Omitting the extraction step will, for many s les, be the key to success if there is limited s le DNA. DNA profiles were generated by direct PCR from 160 fingermarks after treatment with one of the following dactyloscopic fingerprint powders: white hadonite silver aluminium HiFi Volcano silk black or black magnetic fingerprint powder. This was achieved by a combination of an optimised double-swabbing technique and swab media, omission of the extraction step to minimise loss of critical low-template DNA, and additional AmpliTaq Gold
Successful DNA typing of a drug positive urine sample from a race horse
Publisher: Elsevier BV
Date: 11-2007
Bottom‐up in situ proteomic differentiation of human and non‐human haemoglobins for forensic purposes by matrix‐assisted laser desorption/ionization time‐of‐flight tandem mass spectrometry
Publisher: Wiley
Date: 11-10-2017
DOI: 10.1002/RCM.7986
Abstract: The detection and identification of human blood on crime-related items are of particular relevance to many investigations because shed blood can provide evidence of violent contact between in iduals. However, for any detection and identification technique, specificity is a critical performance characteristic to assess that is, whether the technique has the capability to differentiate between human blood (which usually is of relevance to a criminal investigation) and non-human blood (which usually would not be associated with a crime but may be detected incidentally). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) approaches using "top-down" (detection of intact proteins) and "bottom-up" (detection of tryptic peptide markers) were used to detect and identify haemoglobin in blood from humans and from a range of Australian native mammals the technique could be carried out directly on blood stains without the need to extract proteins (i.e., in situ measurement). Imaging of haemoglobin was achieved in bloodied fingermarks, including those that had been enhanced using two "industry standard" fingermark enhancement processes. Differentiation of intact haemoglobin proteins in human and non-human blood using "top-down" MALDI-TOF-MS was difficult. However, in situ "bottom-up" approaches using tandem mass spectrometry (MS/MS) and de novo sequencing of tryptic digest peptides allowed unambiguous differentiation. Imaging mass spectrometry of human haemoglobin, even when it was mixed with animal blood, was achieved in bloodied fingermarks that had been enhanced using two common processes (staining with Amido Black or dusted with magnetic powder) and "lifted" using adhesive tape. The MALDI-TOF-MS-based in situ "bottom-up" proteomic methodology described here shows great promise for the detection of human blood and even imaging of blood in bloodied fingermarks. The approach is sensitive, can differentiate between human blood and that from many animals (including several Australian native animals), and can be implemented after traditional crime scene fingermark enhancement techniques have been carried out.
The benefits and limitations of expanded Y-chromosome short tandem repeat (Y-STR) loci
Publisher: Elsevier BV
Date: 12-2015
DNA profiles from fingernails using direct PCR
Publisher: Springer Science and Business Media LLC
Date: 13-11-2014
DOI: 10.1007/S12024-014-9626-8
Abstract: We report on the successful routine lification of DNA profiles from small sections of fingernails using direct PCR. The data are from 40 nail clippings from eight donors where approximately 4 mm(2) of nail is added directly to the PCR. The NGM™ kit was used that lifies 15 STR loci plus amelogenin. No increase in cycle number was used and no enrichment of the PCR products was performed. Full DNA profiles were observed in 17 of the 40 profiles with 21 generating partial DNA profiles. The process omits the DNA extraction process, and hence there is no opportunity to quantify the DNA prior to lifying the STRs, but by not performing a DNA extraction step, the amount of DNA available for PCR is maximized. Single source DNA profiles were observed in 29 of the 38 profiles obtained. The source of the DNA is assumed to be adhering to the underside of the nail. This simple method offers a significant reduction in time to generate DNA profiles from nail clippings, such as those taken from victims of mass disasters, and should be included into a forensic process relatively easily as it requires no change to manufacturer's instructions for lification.
Characterisation of novel and rare Y-chromosome short tandem repeat alleles in self-declared South Australian Aboriginal database
Publisher: Springer Science and Business Media LLC
Date: 19-09-2013
DOI: 10.1007/S00414-013-0911-Y
Abstract: Y-chromosome short tandem repeats (Y-STRs) are used in forensic science laboratories all over the world, as their application is wide and often vital in solving casework. Analysis of an in-house database of South Australian self-declared Aboriginal males held by Forensic Science South Australia (FSSA) using the Applied Biosystem's AmpFℓSTR® Yfiler™ PCR Amplification Kit revealed 43 variant Y-STR alleles at 6 of the 17 loci. All variant alleles were sequenced to determine the exact repeat structure for each. As a high level of admixture has previously been found within the SA Aboriginal database, s les were haplogrouped using Y-SNPs to determine their likely geographical origin. Although a number of variant alleles were associated with non-Aboriginal Y-haplogroups, a high frequency was observed within the Australian K-M9 lineage. Detailed knowledge of these variant alleles may have further application in the development of new DNA markers for identification purposes, and in population and evolutionary studies of Australian Aborigines.
Monitoring cell loss through repetitive deposition
Publisher: Wiley
Date: 15-09-2022
Abstract: Touch DNA deposited on items can be visualised by using fluorescent nucleic acid staining dyes. It might be expected that if a person contacts items multiple times, then at each contact fewer cells should be transferred and deposited. Here we report on the use of Diamond Dye (DD) to monitor any reduction in cellular deposition during multiple contacts. A volunteer, who was assigned as a heavy shedder, was asked to deposit a thumbprint using both left and right thumbs for 15 s onto separate clean glass slides. Thumbprints were collected in triplicate with each mark made for 15 s. Immediately after deposition, a second and then third mark was made in the same way. The three consecutive depositions were repeated 30 times to create 90 tested thumbprints. The number of cells within each entire thumbprint was scored using a cell‐counting program, developed in‐house. The number of cells deposited by the second and third depositions showed a decrease in the cell number compared to the first deposition, by ~70% and ~85% respectively. There was no difference between the percentage of cell persistence from whole thumbprints (16 frames) compared to only scoring part of thumbprints (4 frames). The data obtained provide insight into how cells are deposited by touch and how many cells remain on the thumb for subsequent contact events. Such information may account for cell deposition when performing actions (e.g. loading a firearm or repetitively opening a closing zip‐lock bag).
Detection of DNA within fingermarks
Publisher: Elsevier BV
Date: 2013
A technique for the quantification of human and non-human mammalian mitochondrial DNA copy number in forensic and other mixtures
Publisher: Elsevier BV
Date: 09-2008
DOI: 10.1016/J.FSIGEN.2008.03.002
Abstract: The number of mitochondria per cell varies by cell type and the number of mitochondrial DNA (mtDNA) genomes varies per mitochondrion. Biological s les from unknown species are encountered frequently in forensic science investigations and are often contaminated with human mtDNA making analysis difficult. Currently, no techniques to quantify non-human mtDNA are available. We report on a method to accurately quantify, sensitive to 100 copies (1.7fg), mtDNA from human and non-human sources when present as a mixture. The test developed uses the cytochrome b (cytb) and the ribosomal 12S genes on the mitochondrial genome. Universal and human specific fragments of similar size are lified and quantified using SYBR Green. We validate the test with 24 human s les and 27 non-human mammalian s les. The human fraction of a s le can then be subtracted from the universal fraction for an accurate estimation of non-human mtDNA copy number.
Low-cost direct PCR for aged and processed wildlife sample analysis
Publisher: Elsevier BV
Date: 2013
Successful direct STR amplification of hair follicles after nuclear staining
Publisher: Elsevier BV
Date: 12-2015
A novel forensic DNA profiling technique for protected species
Publisher: Elsevier BV
Date: 12-2015
Molecular identification of python species: Development and validation of a novel assay for forensic investigations
Publisher: Elsevier BV
Date: 05-2015
DOI: 10.1016/J.FSIGEN.2014.12.002
Abstract: Python snake species are often encountered in illegal activities and the question of species identity can be pertinent to such criminal investigations. Morphological identification of species of pythons can be confounded by many issues and molecular examination by DNA analysis can provide an alternative and objective means of identification. Our paper reports on the development and validation of a PCR primer pair that lifies a segment of the mitochondrial cytochrome b gene that has been suggested previously as a good candidate locus for differentiating python species. We used this DNA region to perform species identification of pythons, even when the template DNA was of poor quality, as might be the case with forensic evidentiary items. Validation tests are presented to demonstrate the characteristics of the assay. Tests involved the cross-species lification of this marker in non-target species, minimum amount of DNA template required, effects of degradation on product lification and a blind trial to simulate a casework scenario that provided 100% correct identity. Our results demonstrate that this assay performs reliably and robustly on pythons and can be applied directly to forensic investigations where the presence of a species of python is in question.
Visualising latent DNA on swabs
Publisher: Elsevier BV
Date: 10-2018
DOI: 10.1016/J.FORSCIINT.2018.08.016
Abstract: Collection for touch DNA either at scenes or on items submitted to a forensic laboratory is based on assumptions as to where a person made direct contact. In many instances a swab may be applied to an area where no contact has been made. Many swabs may therefore be submitted for DNA profiling on which no DNA is present, resulting in the loss of both time and resources by analysing such swabs. This study has developed a simple, fast, DNA-staining and fluorescence microscopy-based screening method for swabs to indicate if there is any DNA from which to generate a profile. Ten different types of swabs were tested covering the major types used (foam, cotton and nylon). Each swab was treated by: no addition of dye or DNA, addition of dye only, addition of known DNA and addition of dye and DNA. The stain used was Diamond™ Nucleic Acid Dye (DD) and fluorescence microscopy was achieved with a digital microscope equipped with a blue LED light source (480nm) for excitation and an emission filter of 510nm. Two types of s les were tested, either buccal swabs or swabs collected from areas touched by volunteers and all analyses were performed in triplicate. The s les were collected and retained at room temperature with time intervals of 0 day, 7 days, 14 days, 21 days, and 28 days before detection using DD staining and fluorescence microscopy. Seven of the swab types used were found to be unsuitable due to the lack of any difference in the fluorescence detected when no DNA, or only the dye, or a combination of DNA and dye were added. Three swab types (black cotton swab, Ultrafine dental applicator, and Cylinder dental applicator) were found to be much more effective for collection of DNA. Further, stained cellular material retained its fluorescence for up to 4 weeks and swabs containing cellular material that had been stored for four weeks could be stained and visualised. Additionally, DD did not affect DNA profiling. This screening method has the potential to be a routine step in a forensic laboratory to save costs of processing s les where swabs are devoid of any DNA. This technique is rapid, easy, cheap, non-destructive and safe.
Tiger species identification based on molecular approach
Publisher: Elsevier BV
Date: 12-2009
A novel approach for rapid cell assessment to estimate DNA recovery from human bone tissue
Publisher: Springer Science and Business Media LLC
Date: 11-10-2021
DOI: 10.1007/S12024-021-00428-3
Abstract: We report on the use of a DNA staining dye to locate and record nucleated osteocytes and other bone-related cells within sections of archived formalin-fixed and paraffin-embedded human tibia from which informative DNA profiles were obtained. Eleven of these archived tibia s les were sectioned at a thickness of 5 µm. Diamond™ Nucleic Acid Dye was applied to the sections and cells within the matrix of the bone fluoresced so that their location and number of cells could be photographed. DNA was isolated from these 11 s les using a standard extraction process and the yields were quantified by real-time PCR. Complete STR profiles were generated from ten bone extracts where low-level inhibition was recorded with an incomplete STR profile obtained from one s le with higher inhibition. The stained image of this s le showed that few cells were present. There was a significant relationship between the number of DD-stained cells and the number of alleles obtained (p < 0.05). Staining cells to determine the prevalence of bone cell nuclei allows a triage of s les prior to any subsequent DNA profiling.
Direct PCR Improves the Recovery of DNA from Various Substrates
Publisher: Wiley
Date: 12-08-2015
Abstract: This study reports on the comparison of a standard extraction process with the direct PCR approach of processing low-level DNA swabs typical in forensic investigations. Varying concentrations of control DNA were deposited onto three commonly encountered substrates, brass, plastic, and glass, left to dry, and swabbed using premoistened DNA-free nylon FLOQswabs(™) . Swabs (n = 90) were either processed using the DNA IQ(™) kit or, for direct PCR, swab fibers (~2 mm(2) ) were added directly to the PCR with no prior extraction. A significant increase in the height of the alleles (p < 0.005) was observed when using the direct PCR approach over the extraction methodology when controlling for surface type and mass of DNA deposited. The findings indicate the potential use of direct PCR for increasing the PCR product obtained from low-template DNA s les in addition to minimizing contamination and saving resources.
DNA profiles from fingermarks: A mock case study
Publisher: Elsevier BV
Date: 12-2015
The complete mitochondrial genome of Axis porcinus (Mammalia: Cervidae) from Victoria, Australia, using MiSeq sequencing
Publisher: Informa UK Limited
Date: 26-07-2017
A complementary forensic ‘proteo-genomic’ approach for the direct identification of biological fluid traces under fingernails
Publisher: Springer Science and Business Media LLC
Date: 12-07-2018
DOI: 10.1007/S00216-018-1223-3
Abstract: Violent contact between in iduals during a crime can result in body fluids becoming trapped under the fingernails of the in iduals involved. The traces under fingernails represent valuable forensic evidence because DNA profiling can indicate from whom the trace originated and proteomic methods can be used to determine the type of fluid in the trace, thus providing evidence as to the circumstances surrounding the crime. Here, we present an initial study of an analytical strategy that involves two complementary techniques, direct PCR DNA profiling and direct mass spectrometry-based protein biomarker detection, for the comprehensive examination of traces of biological fluids gathered from underneath fingernails. With regard to protein biomarker detection, direct MALDI-ToF MS/MS is very sensitive, allowing results to be obtained from biological material present on only a few fibres plucked from a microswab used to collect the traces. Human cornulin, a protein biomarker for vaginal fluid, could be detected up to 5 h after it had been deposited under fingernails whereas haemoglobin, a biomarker for blood, is somewhat more persistent under fingernails and could be detected up to 18 h post-deposition. Bottom-up tandem mass spectrometry techniques were used to provide a high level of confidence in assigning the identity of protein biomarkers. nLC-ESI-qToF MS/MS offered higher levels of confidence and the ability to detect traces that had been present under fingernails for longer periods of time, but this performance came with the cost of longer analysis time and a more laborious s ling approach. Graphical abstract ᅟ.
DNA profiles from matchsticks
Publisher: Informa UK Limited
Date: 22-01-2019
Generation of DNA profiles from fabrics without DNA extraction
Publisher: Elsevier BV
Date: 02-2010
DOI: 10.1016/J.FSIGEN.2009.07.006
Abstract: DNA profiles can be obtained from fabrics where a person has made direct contact with clothing. A standard approach is to cut out a section of the fabric and then use a commercially available method to extract and isolate the DNA. Alternative methods to isolate DNA include the use of adhesive tape to remove traces of cellular material from the fabric prior to extraction. We report on a process to obtain full DNA profiles using direct lification from a range of fabrics. The absence of an extraction step both reduces the opportunity for contamination and reduces the loss of DNA during the extraction process, increasing the sensitivity of the process of generating a DNA profile. The process does not require the use of commercially available extraction kits thus reducing the cost of generating a DNA profile from trace amounts of starting material. The results are in part dependent upon the nature of the fabric used to which the DNA has been transferred.
DNA profiling from human bone cells in the absence of decalcification and DNA extraction
Publisher: Wiley
Date: 24-03-2022
Abstract: Bone cells are a suitable substrate for DNA analysis if required to identify the person from whom a s le was taken. Osteocytes, the most abundant cell type in bone, are embedded within mineralized bone matrix. To release DNA from osteocytes for subsequent analyses, either demineralization of the mineral matrix or an overnight incubation is routinely carried out. In this study, we report on a simplified and rapid approach to analyze preserved bone s les that omits this lengthy decalcification process. Nine tibial bone s les were processed to release matrix‐free bone cells after fragmentation without the use of liquid nitrogen. Cell morphology was assessed by microscopy at 220× magnification following staining with Diamond ™ Nucleic Acid Dye. Based on the presence of stained nuclei, s les were processed either using a DNA extraction process or by a semi‐direct PCR process. The analysis of the quantity and quality of DNA isolated by both methods was carried out by real‐time PCR and STR profiling to assess inhibition of PCR and DNA degradation. All s les resulted in informative STR profiles with minimal indication of inhibitors. These results demonstrate a potential approach of STR profiling from matrix‐free bone cells within 8 hours without decalcification and DNA extraction.
An overview to the investigative approach to species testing in wildlife forensic science
Publisher: Springer Science and Business Media LLC
Date: 2011
Shedding light on shedders
Publisher: Elsevier BV
Date: 09-2018
DOI: 10.1016/J.FSIGEN.2018.06.004
Abstract: All previous examinations of the shedder status of in iduals have been based on conclusions inferred from the amount of DNA deposited by donors after they have held an object for a fixed period of time. In all interpretations of shedder status experiments have involved a range uncertainties, especially in regards to results arising from studies carried out in different laboratories. These apply to the efficiency of the swab collecting DNA from the item touched, the amount of DNA left on the swab after attempts to recover it, and the percentage loss of DNA during the lysis and extraction processes. No previous study has attempted to mitigate these uncertainties or verify how much of the DNA deposited was collected through swabbing, how much DNA present on the swab was recovered or how much DNA is lost during the extraction process. We present a study that accurately measures the deposition, collection and lification of DNA deposited by a range of donors allowing for an accurate determination of the shedder status of in iduals. Eleven donors were asked to wash their hands and then deposit a thumbprint onto glass slides by making pressure for 15 seconds 0, 15, 60 and 180 minutes after handwashing. Both left and right thumbs were used and all testing was performed in triplicate. Measurement of the quantity of cellular material deposited on the slides was carried out using DiamondTM Nucleic Acid Dye and fluorescence microscopy on each of 264 thumbprints. Fluorescence microscopy was then used to demonstrate that all the DNA present on the slides was recovered by the swabbing operations and then direct PCR, using the Identifiler™ Plus kit, was used to ensure that none of the DNA present on swabs was lost during DNA profiling. The combination of using a DNA binding dye and direct PCR allowed an accurate means of measuring the extent to which in iduals exhibit different extents of shedding. This small study, 11 donors, showed that in iduals fell into one of three distinct groups: heavy, intermediate, and light shedders, regardless of the hand used.
A mass spectrometry-based forensic toolbox for imaging and detecting biological fluid evidence in finger marks and fingernail scrapings
Publisher: Springer Science and Business Media LLC
Date: 27-04-2017
DOI: 10.1007/S00414-017-1587-5
Abstract: During a crime, biological material such as blood or vaginal fluid may become smeared on the fingers of the victim or suspect or trapped under their fingernails. The type of trapped fluid is extremely valuable forensic information. Furthermore, if either person touches an object at the crime scene with their 'contaminated' finger then a 'contaminated' finger mark may be deposited. Such marks have great value as they could identify not only who deposited the mark but also who they touched and which part of the body they touched. Here, we describe preliminary work towards a 'toolbox' of techniques based on mass spectrometry (MS) for the identification of biological fluid traces under fingernails or the imaging of them in finger marks. Liquid chromatography-multidimensional MS was effective for the detection of protein biomarkers characteristic of vaginal fluid and blood trapped under fingernails, even after hands had been washed. In regard to examination of finger marks for the presence of biological fluids, the most practical implementation of any technique is to integrate it with, but after, routine crime scene finger mark enhancement has been applied. Here, we demonstrate the usage of matrix-assisted laser desorption ionization-time of flight-MS for the detection and mapping of proteins and peptides from body fluids in finger marks, including marks enhanced using aluminium-containing magnetic powder and then 'lifted' with adhesive tape. Hitherto, only small molecules have been detected in enhanced, lifted marks. In a novel development, aluminium in the enhancement powder assisted ionization of small molecules in finger marks to the extent that conventional matrix was not required for MS.
Current Issues with the Investigation of Wildlife Crime in Australia: Problems and Opportunities for Improvement
Publisher: Informa UK Limited
Date: 03-07-2015
Genetic profiling from challenging samples: Direct PCR of touch DNA
Publisher: Elsevier BV
Date: 2013
Related Organisations
University Of Strathclyde
Location: United Kingdom of Great Britain and Northern Ireland
University Of Sussex
Location: United Kingdom of Great Britain and Northern Ireland
Related Funding Activities
Linkage Projects - Grant ID: LP170100889
Start Date: 12-2018
End Date: 04-2022
Amount: $106,705.00
Funder: Australian Research Council
View Funded ActivityLinkage Infrastructure, Equipment And Facilities - Grant ID: LE130100115
Start Date: 2013
End Date: 12-2013
Amount: $180,000.00
Funder: Australian Research Council
View Funded ActivityLinkage Infrastructure, Equipment And Facilities - Grant ID: LE160100154
Start Date: 05-2016
End Date: 12-2017
Amount: $250,000.00
Funder: Australian Research Council
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