Development of DNA aptamers against Treponema denticola

Abstract

The oral spirochete bacterium Treponema denticola is implicated in the etiology of periodontal disease. However, the elucidation of more specific disease correlations is hindered by the lack of reliable and discriminatory diagnostic methods for T. denticola cell detection and enumeration. Aptamers are an emerging class of molecular recognition agents, which have binding properties analogous to antibodies. Their potential application for oral microbial cell detection remains to be firmly established. Objective:

To identify DNA aptamers against a cell-surface protein from Treponema denticola.

Method:

A recombinant form of an established cell surface protein from T. denticola ATCC 35405 was used as the target for the identification of single stranded DNA aptamers. A SELEX approach was used for aptamer selection from a starting pool of ca. 10(e15) 80-mer oligonucleotides containing a central region of 40 nucleotides of random sequence. The sequences of ca. 30 cloned aptamers selected after the final SELEX selection round were determined. The protein-binding affinities of seven aptamer candidates were evaluated using a variety of biophysical methods.

Result:

The bioinformatic analysis of aptamer sequences indicated the presence of various aptamer structures, including G-quadruplex and (TA-rich) non G-quadruplex forms. Results from ELISA-type binding assays revealed that three aptamers specifically bound to the T. denticola protein target with nanomolar affinities. Additional experiments are ongoing.

Conclusion:

Our preliminary results indicate that two aptamers have significant potential to be used as molecular diagnostic agents for the selective identification of T. denticola strain lineages

This abstract is based on research that was funded entirely or partially by an outside source: Research Grants Council of Hong Kong, General Research Fund (#781911)