scholarly journals Optimisation of rapid STR analysis using a standard DNA forensic pipeline

2019 ◽  
Author(s):  
Katharine Gammon ◽  
Carl Mayers
Keyword(s):  
Capillary Electrophoresis ◽  
Pcr Amplification ◽  
Buccal Cell ◽  
Forensic Dna ◽  
Direct Pcr ◽  
Str Analysis ◽  
Dna Profile ◽  
Fta Cards ◽  
Dna Profiles ◽  
Reference Samples

Previous studies in published literature have reported on various alterations to STR mastermixes, protocols and instrumentation in order to reduce the time taken to generate forensic DNA profiles from reference and casework type samples. In this study, we demonstrate how altering default PCR amplification and capillary electrophoresis protocols in our existing DNA profiling pipeline can reduce the overall time taken to generate a DNA profile from buccal cell reference samples. GlobalFiler Express STR mastermix was used with direct PCR from FTA cards, run on altered PCR protocols and CE settings, and results compared to the standard evaluated settings used in our laboratories. This study demonstrated that full DNA profiles could be recovered in less than 80 minutes in comparison to our standard time of 97 – 102 minutes whilst utilising existing reagent kits and instrumentation, with only minor modifications to protocols.

scholarly journals Direct PCR amplification from saliva sample using non-direct multiplex STR kits for forensic DNA typing

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Pankaj Shrivastava ◽  
Toshi Jain ◽  
R. K. Kumawat
Keyword(s):  
Dna Typing ◽  
Pcr Amplification ◽  
Turnaround Time ◽  
Forensic Dna ◽  
Direct Pcr ◽  
Forensic Dna Typing ◽  
Dna Profile ◽  
Pre Treatment ◽  
Dna Profiles ◽  
Direct Amplification

AbstractDue to its proficiency to provide the most discriminating results for forensic applications, medical research and anthropological studies, multiplex PCR based STR analysis has been established as the most efficient technique in the forensic DNA analysis. Several multiplex amplification kits based on 4, 5 and 6 dyes chemistry are commercially available and used in forensic DNA typing across the globe. These multiplex PCR systems are routinely used for amplification of multiple STR loci (Autosomal, Y and/or X STR’s) in the DNA extracted from various biological samples. In the routine forensic DNA testing, DNA profile obtained is compared with the DNA profile of the reference sample, which takes a certain turnaround time and employs costly lab resources. Successive development in forensic DNA typing have resulted in advent of improved multiplex kits which have reduced the effective analysis time, cost and minimized the number of steps required in comparison to conventional forensic DNA typing. Specialized direct amplification compatible multiplex kits are also available nowadays. These kits are relatively costlier but still require few pre-processing steps, which does not make them worth the hefty cost. Herein, this study, we have used non-direct multiplex STR kits to assess their efficacy for direct amplification. In the present study, 103 saliva samples were directly amplified without any pre-treatment of the samples using thirteen non-direct multiplex kits (4 dyes, 5 dyes and 6 dyes chemistry based) for forensic DNA typing. Here, we report a validated direct PCR amplification protocol from the reference saliva samples by omitting DNA extraction and quantification steps, which resulted in 80% reduction of the turnaround time. The developed protocol is cost effective, time efficient and it does not compromise with the quality of DNA profiles. To the best of our knowledge, this is the first report for direct amplification of DNA with the most commonly used non-direct multiplex STR kits without any pre-treatment of the sample. Complete DNA profiles matching all the essential quality parameters were obtained successfully from all the tested samples.

scholarly journals DMSO Improves the Ski-Slope Effect in Direct PCR

2021 ◽  
Vol 11 (4) ◽  
pp. 1943
Author(s):  
Joo-Young Kim ◽  
Ju Yeon Jung ◽  
Da-Hye Kim ◽  
Seohyun Moon ◽  
Won-Hae Lee ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Pcr Amplification ◽  
Analytical Techniques ◽  
Direct Pcr ◽  
Dna Profile ◽  
Novel Technologies ◽  
Specific Amplification ◽  
Polymerase Chain ◽  
Dna Profiles

Analytical techniques such as DNA profiling are widely used in various fields, including forensic science, and novel technologies such as direct polymerase chain reaction (PCR) amplification are continuously being developed in order to acquire DNA profiles efficiently. However, non-specific amplification may occur depending on the quality of the crime scene evidence and amplification methods employed. In particular, the ski-slope effect observed in direct PCR amplification has led to inaccurate interpretations of the DNA profile results. In this study, we aimed to reduce the ski-slope effect by using dimethyl sulfoxide (DMSO) in direct PCR. We confirmed that DMSO (3.75%, v/v) increased the amplification yield of large-sized DNA sequences more than that of small-sized ones. Using 50 Korean buccal samples, we further demonstrated that DMSO reduced the ski-slope effect in direct PCR. These results suggest that the experimental method developed in this study is suitable for direct PCR and may help to successfully obtain DNA profiles from various types of evidence at crime scenes.

scholarly journals Assessment of QIAGEN™ Investigator® 24plex GO! kit workflow for autosomal STR profiling of forensic reference samples

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Hashom Mohd Hakim ◽  
Hussein Omar Khan ◽  
Siti Afifah Ismail ◽  
Nurul Hazirah Mat Lazim ◽  
Japareng Lalung ◽  
...  
Keyword(s):  
Pcr Amplification ◽  
Dna Profiling ◽  
Buccal Swab ◽  
Forensic Dna ◽  
Str Typing ◽  
Str Analysis ◽  
Dna Database ◽  
Autosomal Str ◽  
Dna Template ◽  
Reference Samples

Abstract Background DNA profiling has proven to be a valuable technique for identification of individuals in crime. Currently, the technique targets several short tandem repeat (STR) regions in human genome. However, increasing number of samples submitted for STR analysis may lead to delays due to the limited number of experienced analysts who might be available at any given moment and the time taken to complete lengthy DNA profiling procedures. This study was conducted to test the specificity, repeatability, reproducibility and robustness of Investigator® 24plex GO! kit for genotyping of reference samples submitted to the Royal Malaysian Police Forensic DNA Laboratory for DNA database. Material and methods In this study, Investigator® 24plex GO! kit was used to directly amplify STR loci from buccal swab cell of reference samples that had previously been STR typed using GlobalFiler™ Express kit. Capillary electrophoresis was carried out on a 3500xL Genetic Analyser using POP-4® Polymer. Amplified products were assigned to particular STR alleles using the GeneMapper ID-X version 1.4 software. Results Our study shows that STR profiles generated using Investigator® 24plex GO! gave concordance results with those previously obtained using the GlobalFiler™ Express kit. In addition, quality sensors included in the kit are of particular importance for determining the effectiveness of the PCR reaction and help to indicate the nature and quantity of DNA template for PCR amplification. Conclusion The Investigator® 24plex GO! kit is reliable for STR typing of reference samples.

scholarly journals Efficient DNA Sampling in Burglary Investigations

Genes ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 26
Author(s):  
Colin Charles Tièche ◽  
Markus Dubach ◽  
Martin Zieger
Keyword(s):  
Crime Scene ◽  
Forensic Dna ◽  
Dna Profile ◽  
Dna Database ◽  
Dna Sampling ◽  
High Incidence ◽  
Crime Scene Investigators ◽  
Dna Collection ◽  
Private Homes ◽  
Reference Samples

In terms of crime scene investigations by means of forensic DNA-analyses, burglaries are the number one mass crime in Switzerland. Around one third of the DNA trace profiles registered in the Swiss DNA database are related to burglaries. However, during the collection of potential DNA traces within someone’s residence after a burglary, it is not known whether the sampled DNA originated from the perpetrator or from an inhabitant of said home. Because of the high incidence of burglaries, crime scene investigators usually do not collect reference samples from all the residents for economical and administrative reasons. Therefore, the presumably high probability that a DNA profile belonging to a person authorized to be at the crime scene ends up being sent to a DNA database for comparison, has to be taken into account. To our knowledge, no investigation has been made to evaluate the percentage of these non-perpetrator profiles straying into DNA databases. To shed light on this question, we collected reference samples from residents who had been victims of recent burglaries in their private homes. By comparing the profiles established from these reference samples with the profiles generated from trace DNA, we can show that the majority of the DNA samples collected in burglary investigations belong to the residents. Despite the limited number of cases included in the study, presumably due to a crime decline caused by the pandemic, we further show that trace DNA collection in the vicinity of the break and entry area, in particular window and door glasses, is most promising for sampling perpetrator instead of inhabitant DNA.

Forensic DNA typing by capillary electrophoresis using the ABI Prism 310 and 3100 genetic analyzers for STR analysis

2004 ◽  
Vol 25 (1011) ◽  
pp. 1397-1412 ◽  
Author(s):  
John M. Butler ◽  
Eric Buel ◽  
Federica Crivellente ◽  
Bruce R. McCord
Keyword(s):  
Capillary Electrophoresis ◽  
Dna Typing ◽  
Forensic Dna ◽  
Forensic Dna Typing ◽  
Str Analysis

scholarly journals Direct PCR amplification of the HVSI region in mitochondrial DNA from buccal cell swabs

2012 ◽  
Vol 64 (3) ◽  
pp. 851-858 ◽  
Author(s):  
Natasa Kovacevic-Grujicic ◽  
S. Davidovic ◽  
Dijana Takic ◽  
Marija Mojsin ◽  
Milena Stevanovic
Keyword(s):  
Mitochondrial Dna ◽  
Dna Analysis ◽  
Pcr Amplification ◽  
Buccal Cell ◽  
Buccal Cells ◽  
Direct Pcr ◽  
Long Term Storage ◽  
Short Period ◽  
Term Storage

Amplification of human mitochondrial DNA (mtDNA) has been widely used in population genetics, human evolutionary and molecular anthropology studies. mtDNA hypervariable segments I and II (HVSI and HVSII) were shown to be a suitable tool in genetic analyses due to the unique properties of mtDNA, such as the lack of recombination, maternal mode of inheritance, rapid evolutionary rate and high population-specific polymorphisms. Here we present a rapid and low-cost method for direct PCR amplification of a 330 bp fragment of HVSI from buccal cell samples. Avoiding the DNA isolation step makes this method appropriate for the analysis of a large number of samples in a short period of time. Since the transportation of samples and fieldwork conditions can affect the quality of samples and subsequent DNA analysis, we tested the effects of long-term storage of buccal cell swabs on the suitability of such samples for direct PCR amplification. We efficiently amplified a 330 bp fragment of HVSI even after the long-term storage of buccal cells at room temperature, +4?C or at -20?C, for up to eight months. All examined PCR products were successfully sequenced, regardless of sample storage time and conditions. Our results suggest that the direct PCR amplification of the HVSI region from buccal cells is a method well suited for large-scale mtDNA population studies.

scholarly journals Short tandem repeat typing technologies used in paternity testing: a case study

2020 ◽  
pp. 04-09
Author(s):  
Amany Abdel Hakim Mousa ◽  
Dena Mohamed Naguib Abdel Moawed ◽  
Adel Mohamed Abdallah Elberry
Keyword(s):  
Tandem Repeat ◽  
Short Tandem Repeat ◽  
Nuclear Dna ◽  
Pcr Amplification ◽  
Str Typing ◽  
Biological Father ◽  
Str Analysis ◽  
Dna Profile ◽  
Federal Bureau Of Investigation ◽  
Short Tandem

Background: In forensic field, for establishing the paternity of disputed offspring, STR typing (autosomal, and Y typing) are commonly used to dissolve the cases. Short tandem repeat (STR) technology is used to evaluate specific regions (loci) within nuclear DNA. Variability in STR regions can be used to distinguish one DNA profile from another. The Federal Bureau of Investigation (FBI) uses a standard set of 13 specific STR regions for CODIS. Aim of the study: The present work was done to find out the biological father of child in a case where mother was killed by her husband for the doubt of her pregnancy. Methods: After taking blood samples from the murdered mother, the baby and the killer husband, Autosomal STRs Using AmpFlSTR Identifiler Plus PCR Amplification Kit and Y-STR analysis using AmpFlSTR Yfiler PCR Amplification Kit were done to solve the disputed paternity case. Results: After the analysis of DNA STRs profiles of autosomal and Y-chromosome markers, the dead fetus was proved to be the son of the killer father. Conclusions: Autosomal STR multiplex analysis combined with Y-STR analysis help to reach a conclusion in disputed paternity cases with greater confidence. Keywords: Disputed; Paternity; Multiplex; Short tandem reprats; Y-STR; Analysis

Conditional Genotypic Probabilities for Microsatellite Loci

Genetics ◽  
2000 ◽  
Vol 155 (4) ◽  
pp. 1973-1980
Author(s):  
Jinko Graham ◽  
James Curran ◽  
B S Weir
Keyword(s):  
Short Tandem Repeats ◽  
Tandem Repeats ◽  
Dirichlet Distribution ◽  
Forensic Dna ◽  
Match Probability ◽  
Microsatellite Genotype ◽  
Allelic State ◽  
Forensic Assessments ◽  
Dna Profiles ◽  
Short Tandem

Abstract Modern forensic DNA profiles are constructed using microsatellites, short tandem repeats of 2–5 bases. In the absence of genetic data on a crime-specific subpopulation, one tool for evaluating profile evidence is the match probability. The match probability is the conditional probability that a random person would have the profile of interest given that the suspect has it and that these people are different members of the same subpopulation. One issue in evaluating the match probability is population differentiation, which can induce coancestry among subpopulation members. Forensic assessments that ignore coancestry typically overstate the strength of evidence against the suspect. Theory has been developed to account for coancestry; assumptions include a steady-state population and a mutation model in which the allelic state after a mutation event is independent of the prior state. Under these assumptions, the joint allelic probabilities within a subpopulation may be approximated by the moments of a Dirichlet distribution. We investigate the adequacy of this approximation for profiled loci that mutate according to a generalized stepwise model. Simulations suggest that the Dirichlet theory can still overstate the evidence against a suspect with a common microsatellite genotype. However, Dirichlet-based estimators were less biased than the product-rule estimator, which ignores coancestry.

scholarly journals Autotetraploid Emergence via Somatic Embryogenesis in Vitis vinifera Induces Marked Morphological Changes in Shoots, Mature Leaves, and Stomata

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1336
Author(s):  
Caterina Catalano ◽  
Loredana Abbate ◽  
Antonio Motisi ◽  
Dalila Crucitti ◽  
Vincenzo Cangelosi ◽  
...  
Keyword(s):  
Somatic Embryogenesis ◽  
Morphological Changes ◽  
Ploidy Levels ◽  
Dna Profile ◽  
Morphological Alterations ◽  
Cell Pair ◽  
Mature Leaves ◽  
Induction Process ◽  
Ploidy Changes ◽  
Dna Profiles

Polyploidy plays an important role in plant adaptation to biotic and abiotic stresses. Alterations of the ploidy in grapevine plants regenerated via somatic embryogenesis (SE) may provide a source of genetic variability useful for the improvement of agronomic characteristics of crops. In the grapevine, the SE induction process may cause ploidy changes without alterations in DNA profile. In the present research, tetraploid plants were observed for 9.3% of ‘Frappato’ grapevine somatic embryos regenerated in medium supplemented with the growth regulators β-naphthoxyacetic acid (10 µM) and N6-benzylaminopurine (4.4 µM). Autotetraploid plants regenerated via SE without detectable changes in the DNA profiles were transferred in field conditions to analyze the effect of polyploidization. Different ploidy levels induced several anatomical and morphological changes of the shoots and mature leaves. Alterations have been also observed in stomata. The length and width of stomata of tetraploid leaves were 39.9 and 18.6% higher than diploids, respectively. The chloroplast number per guard cell pair was higher (5.2%) in tetraploid leaves. On the contrary, the stomatal index was markedly decreased (12%) in tetraploid leaves. The observed morphological alterations might be useful traits for breeding of grapevine varieties in a changing environment.

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