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Analysis of nucleic acid amplification products has become the gold standard for applications such as pathogen detection and characterisation of single nucleotide polymorphisms and short tandem repeat sequences. The development of real-time PCR and melting curve analysis using fluorescent probes has simplified nucleic acid analyses. However, the cost of probe synthesis can be prohibitive when developing large panels of tests. We describe an economic two-stage method for probe synthesis, and a new method for nucleic acid sequence analysis which together considerably reduce costs. The analysis method utilises three-strand and four-strand hybridisation complexes for the detection and identification of nucleic acid target sequences by real-time PCR and fluorescence melting.

Original publication




Journal article


Mol Cell Probes

Publication Date





228 - 236


HyBeacon, Melting curve analysis, Oligonucleotide probe synthesis, Real-time PCR, Universal probes, Chlamydia trachomatis, DNA Primers, Fluorescent Dyes, Nucleic Acid Amplification Techniques, Nucleic Acid Hybridization