Sabanayagam, Chandran R., Boston Univ, Boston, United States
Berkey, Cristin, Boston Univ, Boston, United States
Lavi, Uri, Boston Univ, Boston, United States
Cantor, Charles R., Boston Univ, Boston, United States
Smith, Cassandra L., Boston Univ, Boston, United States

We present an assay to detect single-nucleotide polymorphisms on a chip using molecular DNA switches and isothermal rolling-circle amplification. The basic principle behind the switch is an allele-specific oligonucleotide circularization, mediated by DNA ligase. A DNA switch is closed when perfect hybridization between the probe oligonucleotide and target DNA allows ligase to covalently circularize the probe. Mismatches around the ligation site prevent probe circularization, resulting in an open switch. DNA polymerase is then used to preferentially amplify the closed switches, via rolling-circle amplification. The stringency of the molecular switches yields 102-103 fold discrimination between matched and mismatched sequences.