A quantitative approach to detect and overcome PCR inhibition in ancient DNA extracts

Authors: C. King, R. Debruyne, M. Kuch, C. Schwarz, H. Poinar
Journal: BioTechniques, doi:10.2144/000113244

Inhibition is problematic in many applications of PCR, particularly those involving degraded or low amounts of template DNA, when simply diluting the extract is undesirable. Two basic approaches to monitoring inhibition in such samples using real-time or quantitative PCR (qPCR) have been proposed. The first method analyzes the quantification cycle (Cq) deviation of a spiked internal positive control. The second method considers variations in reaction efficiency based on the slopes of individual amplification plots. In combining these methods, we observed increased Cq values together with reduced amplification efficiencies in some samples, as expected; however, deviations from this pattern in other samples support the use of both measurements. Repeat inhibition testing enables optimization of PCR facilitator combinations and sample dilution such that DNA yields and/or quantitative accuracy can be maximized in subsequent PCR runs. Although some trends were apparent within sample types, differences in inhibition levels, optimal reactions conditions, and expected recovery of DNA under these conditions suggest that all samples be routinely tested with this approach.

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Time-Dependency of Molecular Rates in Ancient DNA Datasets, a Sampling Artifact?

Authors: R. Debruyne, H. N. Poinar
Journal: Systematic Biology, doi:10.1093/sysbio/syp028

It is common knowledge that the instantaneous rate of mutation (RoM) in DNA sequences exceeds the long-term rate of substitution (RoS) when measured in interspecific phylogenetic analyses. The neutral theory of molecular evolution describes this temporary excess diversity as transient polymorphisms either removed from the population through the actions of purifying selection or fixed by random genetic drift over a few generations (Kimura 1983). Observations of these “accelerations” in the molecular rates within recent evolutionary time have been documented (Parsons et al. 1997; Lambert et al. 2002); however, they did not resolve the magnitude and duration of this phenomenon. Howell et al. (2003) have addressed these issues through pedigree analyses of human mitochondrial (mt) hypervariable region (HVR) sequences and have suggested a 5- to 10-fold acceleration compared with the long-term RoS. In addition, Burridge et al. (2008) have shown that the calibration of the mt clock for galaxiid fishes using geological divergence dates with cytochrome b and control region sequences supports a transition period during which the RoM would decrease toward the RoS extending up to ~200 kyr (Burridge et al. 2008). However, the general applicability of these specific results remains untested…

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Darwin 2009 celebration in France

Location: Collège de France, Paris, FR
Date: June 10-12, 2009
Participant: Régis Debruyne
Communication: “Le progrès c’est l’histoire des impossibilités réalisées”: le clonage du mammouth est-il en cours?

Régis Debruyne had a recent opportunity to make use of his French at the colloque entitled “Cent Cinquante ans après l’Origine des espèces: du darwinisme de Darwin à l’évolutionnisme contemporain” of the Collège de France where he presented a talk about both the scientific and the mythologic part of resurrecting the woolly mammoth.

You can reach a complete abstract [in French] of this communication on Reg’s blog.

Evoltree Ancient DNA Workshop

Location: Borrowdale, Lake District, UK
Date: May 4-7, 2009
Participant: Régis Debruyne
Communication: Ancient DNA in revolutions and crises

Régis was part of the Evoltree aDNA workshop setup by the European Network Evoltree on the invitation of Prof. Richard Bradshaw (Univ. Liverpool, Dept. of Geography).

Biotechnology Initiative Lectures

Location: University of Toronto, CA
Date: April 15, 2009
Participant: Régis Debruyne
Communication: Ancient DNA and the benefits of Time Travel

Régis Debruyne took part to the Biotechnology Initiative Lectures of the CBERC, which took place in the McLeod auditorium of the University of Toronto.

These lectures, dedicated to highschool students and life Sciences teachers, met an audience of roughly 450 people, and were a great opportunity to bridge the gap between the realm of scientific research and the one of highschool biology.