Checkerboard DNA-DNA Hybridization Technology Using Digoxigenin Detection

Checkerboard DNA-DNA hybridization (CKB) is a technique that provides a simultaneous quantitative analysis of 40 microbial species against up to 28 mixed microbiota samples on a single membrane; using digoxigenin (DIG)-labeled, whole-genome DNA probes. Developed initially to study the predominantly Gram-negative dental plaque microorganisms involved in periodontitis, we modified the probe species composition to focus on putative pathogens involved in the development of dental caries. CKB analysis is applicable to species from other biodiverse ecosystems and to a large number of samples. The major limitations are that high-quality DNA is required for the preparation of DIG-labeled probes and standards, and that probe specificity requires careful evaluation. Overall, CKB analysis provides a powerful ecological fingerprint of highly biodiverse microbiota based on key cultivable bacteria.