Analysis of salivary DNA evidence from a bite mark on a body submerged in water.

A female body was recovered after approximately 5.5 h in a river with slow-moving current. On the victim's right breast, a patterned injury was discovered and determined to be from human adult teeth. Evidence was collected according to established techniques including recovery of saliva from the bite mark area despite the body being found submerged in water. DNA analysis by PCR using polymorphic STR markers revealed a DNA profile of mixed origin. In addition to the victim's DNA profile, a genotype contribution from the perpetrator was identified as a minor component. The DNA typing results from the bite mark correlated with the DNA typing results obtained from other biological trace evidence identified from the victim's genital samples. The bite mark and the DNA evidence were used to screen suspects and played an important role in obtaining resolution of this case. Consequently, it is advisable that investigators routinely swab for salivary DNA in bite mark cases, even when the amount is thought to be minimal.

Removal of a PCR inhibitor and resolution of DNA STR types in mixed human-canine stains from a five year old case.

The analysis of biological trace evidence from a reopened investigation into a 1991 murder from Vernon, B.C. revealed mixed human and dog bloodstains on blue jean pants that contained a PCR inhibitory substance. The presence of the inhibitory substance was detected by the inhibition caused from adding a small aliquot of the test DNA extract into a PCR reaction designed to produce a known standard product. The removal of the PCR inhibitory substance was accomplished by treating the extracted DNA with Thiopropyl Sepharose 6B beads. DNA profiles from two human contributors and a canine were obtained using species specific polymorphic STR markers. The two human DNA profiles obtained from blue jean pants were resolved, one matched the suspect and the other matched the victim. The DNA profile from the canine component matched that obtained from the known sample of the victim's dog who was also slain during the assault. This evidence along with other DNA typing evidence was critical in obtaining a resolution of the case.

Isolation of a novel G protein-coupled receptor (GPR4) localized to chromosome 19q13.3.

We present the cloning and sequencing of the human gene for a novel G-protein coupled receptor (GPR4), from the critical myotonic dystrophy (DM) region on chromosome 19q13.3. The homologous porcine gene was isolated and sequenced as well. The genes of both species are intronless and contain an open reading frame encoding a protein of 362 amino acids. In human, two isoforms of GPR4 are expressed, differing in their 3' untranslated region due to the use of alternate polyadenylation signals and measuring approximately 2.8 and 1.8 kb, respectively. Northern blot analysis showed that GPR4 is widely expressed, with higher levels in kidney, heart, and especially lung, where it is at least fivefold greater than in other tissues. Sequence analysis suggests that GPR4 is a peptide receptor and shares strongest homologies with purinergic receptors and receptors for angiotensin II, platelet activating factor, thrombin, and bradykinin.

The 1.5-Mb region spanning the myotonic dystrophy locus shows uniform recombination frequency.

The myotonic dystrophy (DM) mutation has been identified as a heritable unstable CTG trinucleotide repeat sequence. The intergenerational amplification of this sequence is an example of a new class of dynamic mutations responsible for human genetic diseases. To ascertain whether recombination activity influences, or is affected by, the presence of this unique sequence, a comprehensive study of the physical and genetic mapping data for the 1.5-Mb region of human chromosome 19q13.3, which contains the DM locus, was conducted. The recombination rate for this region was examined by correlating genetic distance to physical distance for six selected marker loci. The following markers span the DM region: 19qCEN-p alpha 1.4 (D19S37)-APOC2-CKM-pE0.8 (D19S115)-pGB2.6 (DM)-p134c (D19S51)-19qTER. Initial linear regression analysis of these two parameters failed to reveal a significant linear correlation (coefficient of determination, r2 = .19), suggesting nonuniform rates of recombination. However, the presence of a recombination hot spot was believed to be unlikely, as the marker-to-marker pairs that showed the greatest deviation in recombination frequency were not restricted to a specific region of the 1.5 Mb studied and had relatively broad confidence intervals, as reflected by low LOD values. A second linear regression analysis using only marker intervals with high LOD scores (Zmax > 22) showed linear correlation (r2 = .68) for the entire 1.5-Mb region. This analysis indicated a relatively uniform recombination frequency in the 1.5-Mb region spanning the DM locus. Furthermore, the recombinations observed were neither under- nor overrepresented on DM chromosomes. Consequently, recombination activity is unlikely to influence, or be affected by, the presence of the DM mutation.

Hae III--a suitable restriction endonuclease for restriction fragment length polymorphism analysis of biological evidence samples.

Hae III has been selected by our laboratories as the restriction endonuclease of choice for restriction fragment length polymorphism analysis of forensic science samples. The enzyme is compatible with the D2S44 probe system and generates relatively small DNA fragments for that marker system. Similarly, Hae III is compatible with several other independent polymorphic loci, including D1S7, D4S139, D16S85, D17S74, D17S79, D14S13, and D20S15. Hae III is functional under a variety of adverse conditions for DNA digestion and is not affected by the methylation pattern in mammals. Finally, Hae III is a relatively inexpensive restriction endonuclease.

A simple and sensitive method for quantifying human genomic DNA in forensic specimen extracts.

The analysis of DNA restriction fragment length polymorphisms by Southern blot hybridization requires that sufficient quantities of high molecular weight genomic DNA be extracted from biological specimens. Prior to analysis, it is necessary to determine the quantity and quality of the extracted DNA. For many applications, it is also desirable to determine the amount of DNA which is of human origin. In this report, we describe a simple and highly sensitive procedure for the specific quantification of human genomic DNA in forensic extracts or any biological sample. A small fraction of the extract is immobilized onto a nylon membrane and subsequently hybridized to p17H8 (D17Z1), a cloned probe which detects highly repetitive, primate-specific alpha satellite DNA. The procedure requires less than four hours to complete and can be used to quantify subnanogram amounts of hybridizable human genomic DNA.

Bloodstain characterization in the EAP, Hp, Hb, AK and Glo I typing systems using minigels and the PhastSystem.

Application of minigels and the PhastSystem to obtain phenotyping results from bloodstains in the EAP, Hp, AK, and Glo I typing systems was investigated. Nonequilibrium isoelectric focusing with 4-6.5 PhastGel produced readily interpretable phenotypes in the EAP typing system. Both 4-6.5 and 5-8 PhastGel produced AK typing system phenotypes using nonequilibrium isoelectric focusing conditions. The 8-25% PAG PhastGel developed by two staining techniques allowed discrimination of phenotypes for the Hp typing system. Phenotypes from the Glo I typing system were also obtained with this gel type. Variant haemoglobins could be detected on pH 5-8 PhastGel using isoelectric focusing conditions. Much potential for standardized, rapid phenotyping of bloodstains was found to exist utilizing the PhastSystem.