News News

Optimization of protocols for DNA extraction from fecal samples

High-throughput sequencing offers new possibilities in molecular ecology and conservation studies. However, its potential has not yet become fully exploited for noninvasive studies of free–ranging animals, such as those based on feces. High–throughput sequencing allows sequencing of short DNA fragments and could allow simultaneous genotyping of a very large number of samples and markers at a low cost. The application of high throughput genotyping to fecal samples from wildlife has been hindered by several labor intensive steps. Alternative protocols which could allow higher throughput were evaluated for two of these steps: sample collection and DNA extraction. Two different field sampling and seven different DNA extraction methods were tested on grey wolf (Canis lupus) feces. There was high variation in genotyping success rates. The field sampling method based on surface swabbing performed much worse than the extraction from a fecal fragment. In addition, there is a lot of room for improvement in the DNA extraction step. Optimization of protocols can lead to very much more efficient, cheaper and higher throughput noninvasive monitoring. Selection of appropriate markers is still of paramount importance to increase genotyping success. informacion[at] Sarabia et al (2020) Towards high-throughput analyses of fecal samples from wildlife. Animal Biodiver Conserv 43.2: 271–283 Doi 10.32800/abc.2020.43.0271
Average (0 Votes)

Latest News Latest News


Cross-species amplification of microsatellite loci in sub-Antarctic seabirds

Cross-species amplification of microsatellite loci in sub-Antarctic seabirds

Microsatellite loci are ideal for testing hypotheses relating to genetic segregation at fine spatio-temporal scales. They are also conserved among closely related species, making them potentially useful for clarifying interspecific relationships between recently diverged taxa. However, mutations at primer binding sites may lead to increased nonamplification, or disruptions that may result in decreased polymorphism in nontarget species. Furthermore, high mutation rates and constraints on allele size may also with evolutionary time, promote an increase in convergently evolved allele size classes, biasing measures of interspecific genetic differentiation. Here, next-generation sequencing was used to develop microsatellite markers from a shotgun genome sequence of the sub-Antarctic seabird, the thin-billed prion (Pachyptila belcheri), and tested for cross-species amplification in other Pachyptila and related sub-Antarctic species. It was found that heterozygosity decreased and the proportion of nonamplifying loci increased with phylogenetic distance from the target species. Surprisingly, species trees estimated from interspecific FST provided better approximations of mtDNA relationships among the studied species than those estimated using DC, even though FST was more affected by null alleles. A significantly nonlinear second order polynomial relationship between microsatellite and mtDNA distances was observed. Authors propose that the loss of linearity with increasing mtDNA distance stems from an increasing proportion of homoplastic allele size classes that are identical in state, but not identical by descent. Therefore, despite high cross-species amplification success and high polymorphism among the closely related Pachyptila species, we caution against the use of microsatellites in phylogenetic inference among distantly related taxa. informacion[at] Moodley et al (2015) Evolutionary factors affecting the cross-species utility of newly developed microsatellite markers in seabirds Mol Ecol Res DOI: 10.1111/1755-0998.12372