Single primer experiments validating suggested primer-design locations from the iC-Architect software. For each of the 24 single-primer templates, the agarose gel results are shown with calculated PPI and OBV values arranged from poor to good. Two repeat amplifications were performed at two temperatures: 59°C (lanes 1 and 2) and 57°C (lanes 3 and 4). All amplifications were stopped after 20 cycles, except lane 1, which was stopped at 25 cycles to demonstrate that 20 cycles was sufficient to see the difference in amplification. An internal control amplification was also performed on a portion of the sequence consistent with all templates. Tm values for the single-primers were calculated using the IDT OligoAnalyzer Tool.
Pan et al. BMC Biotechnology 2014 14:10 doi:10.1186/1472-6750-14-10