Detection of NASBA amplified bacterial tmRNA molecules on SLICSel designed microarray probes
1 Dept. of Biotechnology, Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia
2 Estonian Biocentre, Tartu, Estonia
3 Dept. of Bioinformatics, Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia
4 Molecular Diagnostics Research Group, National Centre for Biomedical Engineering Science, National University of Ireland, Galway, Ireland
BMC Biotechnology 2011, 11:17 doi:10.1186/1472-6750-11-17Published: 28 February 2011
We present a comprehensive technological solution for bacterial diagnostics using tmRNA as a marker molecule. A robust probe design algorithm for microbial detection microarray is implemented. The probes were evaluated for specificity and, combined with NASBA (Nucleic Acid Sequence Based Amplification) amplification, for sensitivity.
We developed a new web-based program SLICSel for the design of hybridization probes, based on nearest-neighbor thermodynamic modeling. A SLICSel minimum binding energy difference criterion of 4 kcal/mol was sufficient to design of Streptococcus pneumoniae tmRNA specific microarray probes. With lower binding energy difference criteria, additional hybridization specificity tests on the microarray were needed to eliminate non-specific probes. Using SLICSel designed microarray probes and NASBA we were able to detect S. pneumoniae tmRNA from a series of total RNA dilutions equivalent to the RNA content of 0.1-10 CFU.
The described technological solution and both its separate components SLICSel and NASBA-microarray technology independently are applicative for many different areas of microbial diagnostics.