Detection of NASBA amplified bacterial tmRNA molecules on SLICSel designed microarray probes

BMC Biotechnology

Methodology Article

Kadri Toome¹  Ants Kurg²  Sven Parkel²  Ott Scheler²  Lauris Kaplinski³  Priit Palta³  Maido Remm³  Barry Glynn⁴  Majella Maher⁴  Thomas Barry⁴ 

[1] Dept. of Biotechnology, Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia;Dept. of Biotechnology, Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia;Estonian Biocentre, Tartu, Estonia;Estonian Biocentre, Tartu, Estonia;Dept. of Bioinformatics, Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia;Molecular Diagnostics Research Group, National Centre for Biomedical Engineering Science, National University of Ireland, Galway, Ireland;

关键词: Microarray Probe;    Marker Molecule;    Nonspecific Hybridization;    Nucleic Acid Sequence Base Amplification;    Binding Energy Difference;   

DOI  :  10.1186/1472-6750-11-17

 received in 2010-11-12, accepted in 2011-02-28,  发布年份 2011

来源: Springer

【 摘 要 】

BackgroundWe 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.ResultsWe 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.ConclusionsThe described technological solution and both its separate components SLICSel and NASBA-microarray technology independently are applicative for many different areas of microbial diagnostics.

【 授权许可】

CC BY   
© Scheler et al; licensee BioMed Central Ltd. 2011