Theoretical and Experimental DNA Computation (Natural ...

128 5 Physical Implementations
three hours. Detection PCR was carried out as normal and the products run
out alongside the samples from Experiment 18.
Experiment 20. Exclusion experiment 8
This was a Taq-based exclusion using modified cycling conditions. The ALL
template from Experiment 17 was diluted 1/10 and bound to the Dynabeads,
denatured, washed, and split into three. Three single exclusion reactions were
set up, one each for the v2 colors (in this way the three templates acted as
PCR controls for each other).
Experiment 21. Exclusion experiment 9
Experiment 20 was repeated with modifications to primer extension conditions
aiming to favor the annealing of red primers.
Experiment 22. Multiple tile exclusion
This was the first attempt to exclude more than one tile at a time. It was
thought that expanding the experiment in this way may increase the efficiency
of the exclusion reactions by reducing the overall level of template available
at the detection PCR step.
Experiments 23 and 24. Oligo redesign
New oligos were designed to replace the v3 oligos in the existing chain. By
slotting in oligos with modified characteristics, it was hoped that the principle
of the exclusion technique could be shown to work (even if it was not possible
to execute a full algorithm). The basic design features of the new oligos were
as follows:
• The regions of overlap with v2 and v4 were conserved so that the new
oligos could be incorporated into the old chain.
• The color signatures were increased from six to ten nucleotides in length
to increase stability.
• The three color sequences shared no base homology, i.e., they were different
at each individual base.
• All the oligos were checked for runs of bases, homologies, hairpins, etc.
• Primer Tm were as close as possible to each other (so that primers could
be used in combination under optimal conditions), and all around 60◦C. This was achieved by maintaining a GC ratio of 50% for each primer.
The new sequences are described later in this section.