Rapid and Sensitive PCR-Dipstick DNA Chromatography for Multiplex Analysis of the Oral Microbiota
Year: 2014
Type: article
Abstract: A complex of species has been associated with dental caries under the ecological hypothesis. This study aimed to develop a rapid, sensitive PCR-dipstick DNA chromatography assay that could be read by eye for multiplex and semiquantitative analysis of plaque bacteria. Parallel oligonucleotides were immobilized on a dipstick strip for multiplex analysis of target DNA sequences of the caries-associated bacteria, Streptococcus mutans, Streptococcus sobrinus, Scardovia wiggsiae, Actinomyces species, and Veillonella parvula. Streptavidin-coated blue-colored latex microspheres were to generate signal. Target DNA amplicons with an oligonucleotide-tagged terminus and a biotinylated terminus were coupled with latex beads through a streptavidin-biotin interaction and then hybridized with complementary oligonucleotides on the strip. The accumulation of captured latex beads on the test and control lines produced blue bands, enabling visual detection with the naked eye. The PCR-dipstick DNA chromatography detected quantities as low as 100 pg of DNA amplicons and demonstrated 10- to 1000-fold higher sensitivity than PCR-agarose gel electrophoresis, depending on the target bacterial species. Semiquantification of bacteria was performed by obtaining a series of chromatograms using serial 10-fold dilution of PCR-amplified DNA extracted from dental plaque samples. The assay time was less than 3 h. The semiquantification procedure revealed the relative amounts of each test species in dental plaque samples, indicating that this disposable device has great potential in analysis of microbial composition in the oral cavity and intestinal tract, as well as in point-of-care diagnosis of microbiota-associated diseases.
Source: BioMed Research International
DOI: 10.1155/2014/180323
Language: English
Subfield: Periodontics
Field: Dentistry
Domain: Health Sciences
Open Access status: hybrid
Estimated APC paid: $2,680
Awards: Microbiota profiling of infected root canals using metagenomic analysis, Novel roles of antimicrobials in the process of pulpal regeneration: its relationship with dendritic cells and dental pulp stem cells, Change in infected root canal microflora during the course of root canal therapy, Multifaceted approach to the analyses of dental plaque biofilm targeting dental caries-associated bacteria, and its application to dental caries prevention, Metabolomics of cancer tissue -challenge to the paradoxical energy metabolism of cancer cells, Evaluation of intracanal biota using metagenomics - a new strategy for endodontic treatment, Metabolome analysis of oral biofilm: a possible risk assessment of oral diseases using oral microbiome metabolism, Development of a life enhancement model focusing on dining
OpenAlex ID: W2142555352