Additional File 5.
Figure S3. Optimization of primer design for rhPCR. Design of blocked-cleavable primers for use in rhPCR was optimized using a 103 base synthetic oligonucleotide target. A single unmodified Forward (For) primer was used with different blocked-cleavable Reverse (Rev) primers and compared for their relative ability to prime a PCR assay. Blocked-cleavable Rev primers used the same sequence as the unmodified control Rev primer, with the addition of a rU base, and were serially extended by adding 2, 3, 4, 5, or 6 DNA bases 3'-to the ribonucleotide. All blocked primers ended in a ddC residue. Following 45 cycles of PCR, products were separated by denaturing PAGE, fluorescently stained and visualized by UV excitation. M = oligo size markers (bases).
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Dobosy et al. BMC Biotechnology 2011 11:80 doi:10.1186/1472-6750-11-80