Hypervariable region structure and polymorphism of mtDNA from dental pulp and a family analysis.
Nucleotide sequences of the hypervariable region in the D-loop of mitochondrial DNA (mtDNA) were analyzed using DNA extracted from 140 old dental pulp samples. These sequences were compared with the sequence reported by Anderson et al. Nucleotide substitution in the HV1 region was identified at 77 positions. A C-to-T transition at position 16223 (C16223T) was most frequently detected (77.9%). Fourteen types of C-stretch sequence patterns were detected and the same sequence as Anderson had the highest frequency (57.9%). In the HV2 region, base transitions were identified at 56 positions. A263G was identified in all samples. Seven types of C-stretch were detected, but none had the same sequence as Anderson. In the HV3 region, base transitions were identified at 21 positions. T489C was most frequently identified (64.3%). Five types of C-stretch were detected, and the same sequence as Anderson accounted for 92.9%. The 140 samples were classified into 128 kinds by the sequence patterns of the HV region. Next, using the blood and oral mucosa epithelium from 23 subjects comprising four generations in a family line, the hereditary relationship of mtDNA was examined. All mtDNA types of the first-generation mother were infallibly inherited by the fourth generation. (+info)
Non-cultural detection and molecular genotyping of Neisseria gonorrhoeae from a piece of clothing.
Isolation of Neisseria gonorrhoeae is currently the gold standard for the definitive diagnosis of gonorrhoea and for use in medico-legal cases in the UK. Molecular detection methods are used increasingly but are untested as evidence of infection in a court of law. An isolate of N. gonorrhoeae was obtained from a child and an article of clothing from an adult male who was suspected of sexual abuse of the child. Biochemical and immunological tests were used to confirm the isolate as N. gonorrhoeae. Amplification by PCR using two targets, cppB and ompIII, was used both as further confirmation of the isolate and to detect the presence of gonococcal-specific DNA from the clothing. The relationship of the gonococcal DNA from the child and the adult was investigated using genotyping (N. gonorrhoeae multi-antigen sequence typing; NG-MAST), including a nested PCR for the por gene. Both samples were indistinguishable by NG-MAST and shared the same sequence type, 403. This is the first report of molecular detection and genotyping of N. gonorrhoeae on an article of clothing, which resulted in conviction of the man for sexual assault. (+info)
Forensic DNA and bioinformatics.
The field of forensic science is increasingly based on biomolecular data and many European countries are establishing forensic databases to store DNA profiles of crime scenes of known offenders and apply DNA testing. The field is boosted by statistical and technological advances such as DNA microarray sequencing, TFT biosensors, machine learning algorithms, in particular Bayesian networks, which provide an effective way of evidence organization and inference. The aim of this article is to discuss the state of art potentialities of bioinformatics in forensic DNA science. We also discuss how bioinformatics will address issues related to privacy rights such as those raised from large scale integration of crime, public health and population genetic susceptibility-to-diseases databases. (+info)
Branch migration displacement assay with automated heuristic analysis for discrete DNA length measurement using DNA microarrays.
The analysis of short tandem repeats (STRs) plays an important role in forensic science, human identification, genetic mapping, and disease diagnostics. Traditional STR analysis utilizes gel- or column-based approaches to analyze DNA repeats. Individual STR alleles are separated and distinguished according to fragment length; thus the assay is generally hampered by its low multiplex capacity. However, use of DNA microarray would employ a simple hybridization and detection for field forensics and biology. Here we demonstrate a rapid, highly sensitive method for STR analysis that utilizes DNA microarray technology. We describe two adaptations to accomplish this: the use of competitive hybridization to remove unpaired ssDNA from an array and the use of neural network classification to automate the analysis. The competitive displacement technique mimics the branch migration process that occurs during DNA recombination. Our technique will facilitate the rapid deduction of identity, length, and number of repeats for the multiple STRs in an unknown DNA sample. (+info)
Stable RNA markers for identification of blood and saliva stains revealed from whole genome expression analysis of time-wise degraded samples.
Human body fluids such as blood and saliva represent the most common source of biological material found at a crime scene. Reliable tissue identification in forensic science can reveal significant insights into crime scene reconstruction and can thus contribute toward solving crimes. Limitations of existing presumptive tests for body fluid identification in forensics, which are usually based on chemoluminescence or protein analysis, are expected to be overcome by RNA-based methods, provided that stable RNA markers with tissue-specific expression patterns are available. To generate sets of stable RNA markers for reliable identification of blood and saliva stains we (1) performed whole-genome gene expression analyses on a series of time-wise degraded blood and saliva stain samples using the Affymetrix U133 plus2 GeneChip, (2) consulted expression databases to obtain additional information on tissue specificity, and (3) confirmed expression patterns of the most promising candidate genes by quantitative real-time polymerase chain reaction including additional forensically relevant tissues such as semen and vaginal secretion. Overall, we identified nine stable mRNA markers for blood and five stable mRNA markers for saliva detection showing tissue-specific expression signals in stains aged up to 180 days of age, expectedly older. Although, all of the markers were able to differentiate blood/saliva from semen samples, none of them could differentiate vaginal secretion because of the complex nature of vaginal secretion and the biological similarity of buccal and vaginal mucosa. We propose the use of these 14 stable mRNA markers for identification of blood and saliva stains in future forensic practice. (+info)
Vitreous humour as a potential DNA source for postmortem human identification.
PURPOSE: The aim of this study was the assessment of vitreous humor as a potential DNA for forensic human postmortem identification. MATERIAL AND METHODS: Vitreous humor samples were collected using two alternative approaches from 25 corpses of either sex during autopsies. DNA was extracted by standard organic method. Recovered DNA was quantitiated fluorometrically. AmpFlSTR SGM Plus kit and ABI 310 Genetic Analyzer (Applera) were used to obtain genetic profiles. RESULTS: Different DNA yields were quantitated in vitreous body depending on cause of death and sampling approach. CONCLUSION: Vitreous humor is a potential DNA for forensic human postmortem identification depending on a sampling method used. (+info)
Molecular breeding of polymerases for amplification of ancient DNA.
In the absence of repair, lesions accumulate in DNA. Thus, DNA persisting in specimens of paleontological, archaeological or forensic interest is inevitably damaged. We describe a strategy for the recovery of genetic information from damaged DNA. By molecular breeding of polymerase genes from the genus Thermus (Taq (Thermus aquaticus), Tth (Thermus thermophilus) and Tfl (Thermus flavus)) and compartmentalized self-replication selection, we have evolved polymerases that can extend single, double and even quadruple mismatches, process non-canonical primer-template duplexes and bypass lesions found in ancient DNA, such as hydantoins and abasic sites. Applied to the PCR amplification of 47,000-60,000-year-old cave bear DNA, these outperformed Taq DNA polymerase by up to 150% and yielded amplification products at sample dilutions at which Taq did not. Our results demonstrate that engineered polymerases can expand the recovery of genetic information from Pleistocene specimens and may benefit genetic analysis in paleontology, archeology and forensic medicine. (+info)
DNA analysis of early mediaeval individuals from the Zvonimirovo burial site in Northern Croatia: investigation of kinship relationships by using multiplex system amplification for short tandem repeat loci.
AIM: To perform initial DNA analysis of four selected early mediaeval individuals from the Zvonimirovo burial site in Northern Croatia. METHODS: Investigation of genetic matching of individuals from a "double burial" and of individuals with shared cranial non-metric/metric traits from two single inhumations, located in another block of the cemetery complex, was carried out. DNA from four teeth samples was extracted, quantified, and amplified by polymerase chain reaction (PCR) for short tandem repeat loci, using AmpFlSTR Profiler PCR Amplification Kit. RESULTS: Autosomal STR genotyping generated high parentage probability (PP) as to the matching of the two individuals from the "double burial" site (PP 98.63%), and of two women with shared cranial non-metric/metric traits from neighboring single burials (PP 90.07%). Parentage probability calculations of a possible genetic matching of the subadult from a "double burial" with the adults from single burials 4 and 3, were significantly lower (PP 60.45% and 38.52%). DNA typing for amelogenin confirmed the sex of the three female individuals, estimated previously by morphology. The unknown sex of a subadult was also identified as female. CONCLUSION: Increased parentage probability for autosomal STR loci matches and the presence of a rare allele shared among matched individuals support their possible kinship relationship, in accordance with bioarchaeological data. We assume an intentional double burial based on a close familial relationship, ie two single neighboring inhumations based on consanguinity, rather than a strong social relationship. The kinship lineages remain unknown at this point. (+info)