Validation of a 16-locus fluorescent multiplex system

STR multiplexes have been indispensable for the efficient genotyping of forensic samples. The PowerPlex 16 System contains the coreCODIS loci, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, D21S11, CSF1PO, FGA, THOI, TPOX, vWA, the sex determinant locus, amelogenin, and two pentanucleotide STR loci, Penta D and Penta E. This multiplex satisfies the locus requirements for most national databases and is the most efficient currently available system due to its single PCR amplification. To provide the groundwork for judicial acceptance, including the publication of primer sequences, and to evaluate laboratory-to-laboratory variation, a developmental validation for casework on this commercially available system was performed in 24 laboratories and produced the following conclusions. Amplification was reliable on a variety of thermal cyclers and product could be analyzed on either an ABI PRISM 310 Genetic Analyzer or an ABI PRISM 377 DNA Sequencer. Genotyping using single source samples was consistent between 0.25 and 2 ng of input DNA template with a few laboratories obtaining complete genotypes at 0.0625 ng. However, heterozygote allele imbalance (<60% peak height balance) caused by stochastic effects was observed at a rate of 13% with 0.125 ng DNA and 22% at 0.0625 ng DNA. Mixture analyses were done using a total of 1 ng of DNA template. Most alleles were detected in mixtures of 4 to 1 and some minor alleles were detected in mixtures of 19 to 1. Optimum amplification cycle number was dependent on the sensitivity of the detection instrument used and could also be adjusted to accommodate larger amounts of DNA on solid supports such as FTA paper. Reaction conditions including volume, annealing temperature, and concentrations of primer, AmpliTaq Gold, and magnesium were shown to be optimal yet robust enough to withstand moderate variations without affecting genotype analysis. Environmental, matrix and standard source analyses revealed an ability to obtain complete genotypes in all sample types except those exposed to 80 degrees C for 12-48 days. Finally, comparison of genotype results from the PowerPlex 16 System with other commercially available systems on non-probative reference and forensic samples showed consistent results.     

An improved method for post-PCR purification for mtDNA sequence analysis

Mitochondrial DNA (mtDNA) analysis of forensic samples typically is performed when the quantity and quality of DNA are insufficient for nuclear DNA analysis or when maternal relatives may be the only reference source. Many of the steps required in the analytical process are both lengthy and labor intensive. Therefore, improvements in the process that reduce labor without compromising the quality of the data are desirable. The current procedure requires purification of the amplicons by centrifugal filtration after PCR and prior to cycle sequencing. Because this method requires several manipulations to perform, alternate cleanup procedures were investigated. These include the use of 1) Qiagen QlAquick PCR Purification columns, 2) Concert Rapid PCR Purification columns, and 3) ExoSAP-IT reagent. When the yield of purified amplicons, quality of the sequence profile, and ease of assay were evaluated, the use of ExoSAP-IT reagent for post-amplification purification was chosen to replace the filtration method.     

Crystallization of cerebrospinal fluid in cases of death from ischemic heart disease and ethanol poisoning

Crystallographic analysis of the cerebrospinal fluid (CSF) was carried out in 18 cases of death from coronary disease and 19 cases of death from ethanol poisoning. The crystallograms were evaluated visually and by the stereoscopic picture. Specific features of the CSF crystal colonies growth in subjects dead from the above conditions are determined.     

Population genetics of nine short tandem repeat loci: allele frequency distribution in a Brazilian population sample

Gene and genotype frequencies in relation to the D3S1358, vWA, FGA, TH01, TPOX, CSF1PO, D5S818, D13S317, and D7S820 loci were determined in a sample of 290 unrelated individuals (204 Caucasians and 86 mulattoes) living in the city of S?o Paulo, Brazil. The sex test Amelogenin was also performed in all subjects from our sample, revealing the expected sex in all instances. Allele frequency data obtained from the analysis of these samples were in the usual range of other population groups with similar racial background. In the sample of Caucasian individuals, panmictic proportions were ruled out in relation to TPOX and CSF1PO loci, but only in the latter was the overall frequency of heterozygotes significantly less than expected. In the sample of mulattoes, Hardy-Weinberg proportions were rejected in relation to FGA and CSF1PO loci, but in no instance were the overall numbers of heterozygotes different from the corresponding expected ones under panmixia. Taking into account all this and also the number of tests performed, the degree of genetic heterogeneity of Brazilian populations, and the critical level reached by the significant results (1% < alpha<5%), the departures from panmixia here observed can be considered to be negligible in altering significantly biologic relationship odds calculated under the assumption of random matings.     

A new improved method for extraction of DNA from teeth for the analysis of hypervariable loci

A new method for better recovery of DNA suitable for amplification of hypervariable loci from fragments of teeth, consisting of two steps-scraping and aspiration, and extensive decalcification-is reported. Higher yields of high molecular weight DNA were obtained from the root, pulp, and crown of all kinds of 120 teeth, irrespective of gender, age, and source of teeth. HLA DQA1, 5 poly markers (LDLR, GYPA, HBGG, D7S8, and Gc), and other 12 short tandem repeat loci (HPRTB, F13B, LPL, D13S317, D7S820, D5S818, D21S11, D18S51, FGA, D8S1179, D3S1358, and vWA) could be successfully amplified and typed from recovered DNA.     

Estimation of body exposure to explosion

An ordnance-disposal expert was killed while disposing of a cache of explosives. The likely position of the body was reconstructed by modeling the explosion as an omnidirectional emission of particles from a model of the explosion site and noting the distribution of particles on a model of a human. The applications and limitations of this method in reconstructing the events and correlation with the injuries noted at autopsy are discussed.     

Rethinking the Probative Value of Evidence: Base Rates,Intuitive Profiling,and the “Postdiction” of Behavior

It is argued that American courts may be routinely admitting evidence with little to no probative value and great potential for prejudicial impact. This may be particularly likely with regard to what is essentially intuitive profiling or stereotype related evidence, defined herein as evidence suggesting that the defendant (or other party), or his (her) behavior, fits intuitive profiles (or stereotypes) of the type of person likely to commit the crime or behavior in question. In other words, intuitive profiling evidence is admitted to postdict behavior. Formal empirically based profiling evidence (testimony regarding the fit of a defendant's characteristics or behaviors to formal or scientific profiles of the typical perpetrator of the crime in question for use to prove guilt is inadmissible in American courts. However, we suggest that everyday use of informal intuitive profiles underlies both judicial determinations of probative value diagnosticity, and thus admissibility, of evidence, and jurors' use of the evidence in determining guilt. Demonstrations of the use of base rate information to evaluate the probative value of such intuitive profiling evidence both as evidence of guilt and as evidence of innocence are provided. Demonstrations of both how to evaluate the actual probative value of evidence (when all necessary values are known), and the theoretical limit of its probative value (in circumstances where some values are not known) are provided. It is argued that such evaluations may provide the basis for (1) support of motions to either admit or to exclude evidence, (2) testimony to the jury to help them weigh or interpret evidence, (3) exculpatory profiling (profiling evidence of innocence), (4) pretrial research to establish probative versus prejudicial value of evidence, and (5) sufficiency analyses to determine maximum likelihood of guilt, given multiple items of evidence. Among these, the first two are considered most important, as it can be demonstrated that many profiling characteristics currently admitted in trial (such as evidence of battery to support a murder charge) are not probative of guilt.