Acceptance of domestic cat mitochondrial DNA in a criminal proceeding.

Shed hair from domestic animals readily adheres to clothing and other contact items, providing a source of transfer evidence for criminal investigations. Mitochondrial DNA is often the only option for DNA analysis of shed hair. Human mitochondrial DNA analysis has been accepted in the US court system since 1996. The murder trial of the State of Missouri versus Henry L. Polk, Jr. represents the first legal proceeding where cat mitochondrial DNA analysis was introduced into evidence. The mitochondrial DNA evidence was initially considered inadmissible due to concerns about the cat dataset and the scientific acceptance of the marker. Those concerns were subsequently addressed, and the evidence was deemed admissible. This report reviews the case in regards to the cat biological evidence and its ultimate admission as generally accepted and reliable. Expansion and saturation analysis of the cat mitochondrial DNA control region dataset supported the initial interpretation of the evidence.

Mitochondrial DNA sequence heteroplasmy levels in domestic dog hair.

To assess the level of mtDNA sequence heteroplasmy in dog hair, we sequenced a 612 base pair fragment of the hypervariable region 1 (HVI) in 576 hairs from six dogs representing a range of age, sex, breed, and hair color. Blood and buccal samples were collected from each dog for reference. Three instances of sequence heteroplasmy were observed at nucleotide positions 15627 (G/A), 15628 (T/C) and 15639 (G/A) in two hairs from different dogs. An HVI sequence heteroplasmy frequency of 0.0034 was obtained. The Probability of Identity (PI) value, or probability that two random, unrelated dog hairs share an HVI sequence, and the Power of Discrimination (PD), or probability that two random unrelated dog hairs have different HVI sequences, were determined from the 88 HVI haplotypes represented in the Veterinary Genetics Laboratory database (n=1006) and found to be 0.086 and 0.914, respectively.

Developmental validation of DogFiler, a novel multiplex for canine DNA profiling in forensic casework.

While the analysis of human DNA has been the focus of large-scale collaborative endeavors, non-human forensic DNA analysis has not benefited from the same funding streams and coordination of effort. Consequently, the development of standard marker panels, allelic ladders and allele-specific sequence data comparable to those established for human forensic genetics has lagged. To meet that need for domestic dogs, we investigated sequence data provided by the published 7.6X dog genome for novel short tandem repeat markers that met our criteria for sensitivity, stability, robustness, polymorphic information content, and ease of scoring. Fifteen unlinked tetranucleotide repeat markers were selected from a pool of 3113 candidate markers and assembled with a sex-linked marker into a multiplex capable of generating a full profile with as little as 60pg of nuclear DNA. An accompanying allelic ladder was assembled and sequenced to obtain detailed repeat motif data. Validation was carried out according to SWGDAM guidelines, and the DogFiler panel has been integrated into forensic casework and accepted in courts across the U.S. Applying various formulae for calculating random match probabilities for inbred populations, estimates for this panel of markers have proven to be comparable to those obtained in human forensic genetics. The DogFiler panel and the associated allelic ladder represent the first published non-human profiling system to fully address all SWGDAM recommendations.

Developmental validation of Mini-DogFiler for degraded canine DNA.

Dogs (Canis lupus familiaris) are kept as pets in 39% of American households and are, therefore, a significant source of potentially probative biological evidence. As with any biological evidence, degradation can occur as a consequence of environmental exposure causing fracturing of the DNA and a resulting loss of intact template. Degraded human DNA analysis has benefited from the application of primer sets that amplify shorter nuclear sequences for core STR loci (miniSTRs), resulting in improved DNA profiles. This same approach was applied to our core canine STR loci. The 16-locus "DogFiler" panel was redesigned into three panels of miniSTRs for analysis of degraded canine DNA, with all primer pairs producing amplicons below 205 base pairs in length. These new miniSTR marker panels - known as Mini-DogFiler - were validated according to SWGDAM guidelines, and concordance with the original 16-locus multiplex was demonstrated through genotyping 1244 samples. The combination of these miniSTRs and a half-volume reaction increased the amplification success of degraded and low copy number canine biological samples resulting in a near three-fold increase in reportable alleles. This assemblage of miniSTRs along with the DogFiler panel and associated allelic ladder are the first non-human DNA profiling system to parallel the human forensic paradigm.