Modelling cross‐hybridization on phylogenetic DNA microarrays increases the detection power of closely related species

Abstract

DNA microarrays are a popular technique for the detection of microorganisms. Several approaches using specific oligomers targeting one or a few marker genes for each species have been proposed. Data analysis is usually limited to call a species present when its oligomer exceeds a certain intensity threshold. While this strategy works reasonably well for distantly related species, it does not work well for very closely related species: Cross-hybridization of nontarget DNA prevents a simple identification based on signal intensity. The majority of species of the same genus has a sequence similarity of over 90%. For biodiversity studies down to the species level, it is therefore important to increase the detection power of closely related species. We propose a simple, cost-effective and robust approach for biodiversity studies using DNA microarray technology and demonstrate it on scenedesmacean green algae. The internal transcribed spacer 2 (ITS2) rDNA sequence was chosen as marker because it is suitable to distinguish all eukaryotic species even though parts of it are virtually identical in closely related species. We show that by modelling hybridization behaviour with a matrix algebra approach, we are able to identify closely related species that cannot be distinguished with a threshold on signal intensity. Thus this proof-of-concept study shows that by adding a simple and robust data analysis step to the evaluation of DNA microarrays, species detection can be significantly improved for closely related species with a high sequence similarity.

Phylogenetic Analysis of the Oriental-Palearctic-Afrotropical Members of Anopheles (Culicidae: Diptera) Based on Nuclear rDNA and Mitochondrial DNA Characteristics

“…This finding is in agreement with the view of other researchers indicating that ribosomal gene, including ITS2, are more useful in molecular systematics of various taxa, including Anopheles (42)(43)(44). This locus has also been used in barcoding and DNA array technologies (45,46). Mohanty et al inferred phylogenetic relationships among the species of the subgenus Cellia using the mitochondrial genes COI and COII and the rRNA gene (D3 and ITS2 regions).…”

Molecular characterization of the Indian poultry nodular tapeworm, Raillietina echinobothrida (Cestoda: Cyclophyllidea: Davaineidae) based on rDNA internal transcribed spacer 2 region

“…Furthermore, the ITS2 markers have also been proposed for use in species barcoding and DNA microarrays, thus increasing the detection power of closely related species (Park et al 2007;Engelmann et al 2009). In recent years, several molecular tools including sequence data related to conserved domains have unveiled their utility in molecular identification, taxonomy, systematics and phylogeny of floral, faunal and microbial species, too (Huang et al 2006;Prasad et al 2009a, b;Wiemers et al 2009;Yao et al 2010;Sharma et al 2012).…”

Assessing Macroinvertebrate Biodiversity in Freshwater Ecosystems: Advances and Challenges in DNA-based Approaches

“…Inevitably, as the number of taxa incorporated in an array increases, the scope of these problems becomes increasingly more complex. In addition to the refinement of array design for whole communities, more powerful analytic approaches are needed that maximize the information content from multiple DNA probes in order to identify closely related species (Engelmann et al 2009). In future applications, specifically designed arrays could be used as tools for developing characteristic DNA signatures (Cannon et al 2006), similar to the way in which community typing is currently used for microbial communities through RFLP techniques.…”