Dean P. Hildebrand

Saliva from cheese bite yields DNA profile of burglar: a case report.

Abstract Physical evidence in the form of a high quality bite mark was discovered on a piece of yellow cheese found at the scene of a crime. The cheese had been frozen by police for 10 days after recovery and before submission to the laboratory for testing. The double swab technique was used to collect DNA samples. A sample of the suspect’s blood was obtained. Using PCR-based DNA typing at ten STR loci, (Profiler Plus, Perkin Elmer-Applied Biosystems) it was determined that the DNA from the cheese originated from the suspect. This case illustrates the importance of a) always considering human bite marks as both physical and biological evidence, and b) attempting DNA recovery in any case in which minute traces of saliva may be present, even in situations involving bacteria-rich foods.

Recovery of DNA from Human Teeth by Cryogenic Grinding

DNA has been previously recovered from human teeth for RFLP and PCR-based forensic analysis. In some cases, the maximum amount of undisturbed tooth structure is required for ulterior forensic analysis. But, in most cases, following comprehensive documentation, it is possible to section the tooth longitudinally or horizontally, or crush it to access the DNA-rich core. This technical report describes an alternative method to recover DNA from whole extracted human molar teeth. A 6700 freezer mill was used to pulverize 20 teeth under frozen preparation in liquid nitrogen and sterile conditions. The mean yield of DNA was 30.9 micrograms (18.4 micrograms DNA per gm tooth powder). The resulting fine powder was subjected to organic extraction and subsequently quantified using slot blot hybridization. Aliquots were successfully amplified at three short tandem repeat polymorphic loci. The technique is simple and relatively rapid. Isolation of the samples during pulverization minimizes the risk of contamination.

IDENTIFICATION OF A SKELETON USING DNA FROM TEETH AND A PAP SMEAR

Identification of unknown living or deceased persons using dental treatment records is an established forensic technique. However, some cases remain unidentified, especially when antemortem dental records are not available for comparison to postmortem dental records. Cytological smears have been previously reported to be potential sources of DNA reference samples which can be compared to DNA recovered from found human remains. The case described here involves an adult skeleton which exhibited extensive, complex dental restorative treatment. A putative identification of the found skeleton as a missing woman was established using circumstantial evidence found at the scene. However, it became important to establish a positive identification using reliable scientific methods. When it was discovered that antemortem dental records were not available because the treatment was completed in another country and the treating dentist could not be found, cytological smears stained with Papanicolaou (PAP) stain obtained from the putative decedents medical records were used as a reference DNA sample. DNA was recovered from the teeth of the skeleton using cryogenic grinding. Comparison of the genotypes resulted in the conclusion that the DNA originated from the same source. The use of PAP smears in this way is seen as a valuable resource in cases where positive identification using traditional dental and medical records is not possible.

Spectroscopic and functional studies of a novel quadruple myoglobin variant with increased peroxidase activity

A quadruple variant of horse heart myoglobin (Thr39Ile/Lys45Asp/Phe46Leu/Ile107Phe) that exhibits significantly (approximately 25-fold) greater peroxidase activity than the wild-type protein has been studied to determine its midpoint reduction potential (24(2) mV vs. SHE; pH 6.0, mu = 0.1 M, 25 degrees C) and to characterize the kinetics of its reaction with hydrogen peroxide. In addition, Fourier transform infrared (FTIR) spectra of the carbonyl and azide adducts of the protein have been obtained to gain initial insight into the effects of these substitutions on the ligand binding properties of the reduced and oxidized variant. All of the results obtained in this work are consistent with a variant heme binding pocket with increased hydrophilic character.

Trans effects on cysteine ligation in the proximal HIS93CYS variant of horse heart myoglobin

Three variants of horse heart myoglobin (Mb) in which the proximal His93 residue has been replaced with a Cys residue have been constructed and studied by NMR, EPR, and MCD spectroscopy to evaluate the contributions of proximal and distal residues to the coordination environment of the heme iron in these proteins. Although no experimental conditions were identified that allowed quantitative ligation of the cysteine residue to the heme iron in the His93Cys variant, all of the spectroscopic evidence collected for the His93Cys/His64Ile and His93Cys/His64Val double variants supports the assignment of thiolate as the ligand to iron in the oxidized forms of these variants. The double metMb variants exhibit Soret maxima that are considerably blue-shifted, 1H NMR spectra with decreased mean methyl resonances, and EPR spectra with highly rhombic g values. These spectroscopic data for the Fe(III) variants resemble the corresponding properties reported for ferricytochrome P-450. The decrease in the reduction potential of the double variants by 280 mV relative to wild-type protein is also consistent with the low midpoint potential of cytochrome P-450. MCD spectroscopy of these variants confirms that the proximal cysteine residue is not bound in the reduced forms of these proteins and, in the case of the His93Cys variant, that the distal histidine is coordinated to the iron. Similar coordination environments were created in the ferrimyoglobin variants by cyanogen bromide modification, which resulted in cyanation of the sulfur atom and prevented the ligation of Cys93 to the heme iron.(ABSTRACT TRUNCATED AT 250 WORDS)