Detection and identification of DNA by PCR has opened tremendous possibilities and allows detection of minute quantities of DNA highly specifically. However, PCR remains confined to laboratory settings because of the need of thermocyclers and other analytical equipment. This led to development of isothermal amplification techniques, among which Pad Lock Probe (PLP)-based Rolling Circle Amplification (RCA) has several advantages, but typically also requires a laboratory apparatus of some sort to measure DNA amplification. To circumvent this limitation, while still taking advantage of PLP-based RCA, we developed a colorimetric assay that relies on pH change. Using this assay, we can detect DNA in the low picomolar range and obtain results observable with the naked eye in only 20 min without any requirement for a thermocycler or other complex device, making it a particularly portable assay.
Keywords: Biosensor; Bst DNA polymerase; Colorimetric assay; Padlock probe; Rolling circle amplification; Unbuffered ligation.
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