Multiplex detection of hotspot mutations by rolling circle-enabled universal microarrays

Detection of somatic low abundance mutations in early cancer development re quires a discriminatory, specific, and high-throughput methodology. In this study we report specific, discriminatory detection of low abundance mutati ons through a novel combination of rolling circle amplification (Nat Genet 1998; 19:225-232) and PCR ligation detection reaction on a universal oligon ucleotide microarray (J Mol Biol 1999; 292:251-262). After mutation-specifi c multiplex ligation and hybridization of 17 pairs of probes to a generic m icroarray, the ligated probes were visualized. The multiplex mutation-speci fic ligation is possible only because rolling circle amplification permits quantification of previously undetectable hybridization events conducive to the detection of a single mutation from within a pool of over 100 wild-typ e alleles. This system is readily adaptable to high-throughput automation u sing a robot such as the Biomek platform.