Spotting the haystacks with the right needles

 Published in 2011

Spotting the haystacks with the right needles

Niederstätter,H.; Kralj,P.; Bauer,C.; Parson,W.

PCR is sometimes called the method that turns the proverbial needle in a haystack into a needle stack. Its specificity and sensitivity paved the way for the revolution witnessed in forensic individualization of biological material. STR markers are the “stars amongst the needles” and their multiplexed typing represents the gold standard in human identification, but the analysis of other genetic markers, such as mitochondrial or Y-chromosomal DNA, fills vital niches. The speed and simplicity of these analyses enable the typing of large sample numbers e.g. in population studies or DNA mass screenings. However, testing the entire sample set becomes highly uneconomical when the investigation aims only at a particular part of the sample. Under such framework conditions affordable and reliable pre-screening assays for the high throughput exclusion of e.g. innocents in a DNA dragnet or samples not attributable to the mitochondrial or Y-chromosomal haplogroup under study are desirable to avoid unnecessary STR genotyping or sequencing analyses.

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IDENTIFICATION OF AN ANIMAL CONTRIBUTOR WITHIN MIXTURE TRACES

 Published in 2011

IDENTIFICATION OF AN ANIMAL CONTRIBUTOR WITHIN MIXTURE TRACES BY INTERPRETATION OF THE MITOCHONDRIAL DNA SEQUENCE

Berger,C.; Niederstätter,H.; Steinlechner,M.; Hatzer-Grubwieser,P.; Parson,W.

Species identification of non-human traces is well established in the forensic community and supplements the commonly used human specific DNA analysis. The routinely used Sanger sequencing analysis of the cytochrome b (cytb) gene proved to be one of the most suitable methods to infer the species of an unknown sample over the last years as it is robust and in a lot of cases successful due to the high copy number of mitochondrial (mt) DNA per cell.

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Alternative strategy for whole mitochondrial genome amplification

 Published in 2009

An alternative strategy for whole mitochondrial genome amplification and sequencing suited for lower quality mtDNA

Zimmermann,B.; Huber,G.; Fendt,L.; Parson,W.

Amplification and sequencing of the entire mitochondrial (mt) genome becomes increasingly important in the fields of forensic DNA testing and phylogenetics, as control region sequencing and targeted coding region SNP typing are sometimes not enough for a clear haplogroup assignment. Nowadays the amplification and sequencing of the mtDNA control region (CR) seems to be a routine exercise in some labs, while sequencing of the whole mt genome (~16.6kb) is more demanding, especially when the DNA quality of the samples of interest is low.

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Facile cost-efficient fast and reliable two-stage strategy for high-throughput identification

 Published in 2009

A facile cost-efficient fast and reliable two-stage strategy for the high-throughput identification of samples belonging to mtDNA haplogroup K and its subhaplogroups K1 and K2

Niederstätter,H.; Bauer,C.; Fendt,L.,Parson,W.

We developed for a study focusing on the mitochondrial haplogroup K (hg K) and its subhaplogroups K1 and K2 a two-staged reliable low-cost assay for high throughput identification of samples attributable to these haplogroups in a large Austrian population sample. For this purpose homogeneous allele-specific PCR (the amplification refractory mutation system, ARMS) was used. With hg K respectively non-hg K samples the ARMS specifically amplified either the derived 9055A allele (hg K samples) or the ancestral 9055G allele (non-hg K samples) in the presence of the dsDNA binding fluorescent dye Eva Green. Further subdividing the samples attributed to hg K was performed using an competitive duplex ARMS assay simultaneously interrogating the SNP alleles 1189C (hg K1) and 9716C (hg K2). To differentiate non-hg(K1,K2) or potential K* samples from PCR failure, a short sequence stretch around nucleotide position (ntp) 9055 was co-amplified with the two ARMS products. For both assays, product detection and allele calling were based on the amplicon characteristic melting-point temperatures (T m ) obtained by on-line post-PCR fluorescent dissociation curve analysis (DCA). Both ARMS-DCA assays worked over a broad range of initial DNA concentrations, and enabled the successful analysis of a large population sample without using robotic lab-equipment. We conclude that homogeneous fluorescent ARMS-DCA in general represents a very attractive methodological approach for large-volume studies when the number of SNP markers is limited. It might be also useful for the fast exclusion of the vast majority of specimens in mass-screenings if the haplogroup affiliation of the offender is known.

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Y-chromosome SNP multiplex for haplogroup assignment

 Published in 2009

A Y-chromosome SNP multiplex for haplogroup assignment of West Eurasian men from Tyrol (Austria)

Niederstätter,H.; Erhart,D.; Pitterl,F.; Oberacher,H.; Berger,B.; Parson,W.

Mountain-valleys preset migration routes and define highly structured areas of human settlement. This is likely to result in the formation of complex genetic patterns, reflecting the influence of the landscape’s topology. Y-chromosomal data from studies with high geographic resolution are still in demand. In order to gain a better understanding of the genetic landscape of the male population living in Tyrol, we developed a minisequencing assay for the simultaneous analysis of 19 phylogenetically informative single nucleotide polymorphisms (SNPs) on the human Y-chromosome.

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