US Patent 7977050 Nucleic acid amplification with integrated multiplex detection

A method mediated with in-vitro transcription (“IVT”) which permits miniaturization of multiplexed DNA and RNA analysis, and in which elongation-mediated multiplexed analysis of polymorphisms (eMAP®) is used as the analysis step, is described. Also described is a method mediated with IVT is for selecting a designated strand from T7-tagged double stranded DNA: wherein, the selected strand forms the template for RNA synthesis. In one embodiment, double stranded DNA incorporating the T7 (or other) promoter sequence at the 3′ end or the 5′ end is produced, for example, by amplification of genomic DNA using the Polymerase Chain Reaction (PCR). Also disclosed are nested PCR designs permitting allele analysis in combination with strand selection by IVT. Further, in one embodiment of a homogeneous format for transcription-mediated amplification and multiplexed detection (which may be particularly suited for viral or pathogen detection), encoded microparticles display “looped” capture probe configurations permitting the generation of a signal upon capture of RNA product and real-time assay monitoring.