John M. S. Bartlett.pdf - Bio-Nica.info

86 Hyndman and Mitsuhashi
Fig. 3. Hybridization simulation data.
3.2.2. Hybridization Simulation
The most precise way to view the specificity of a PCR primer is by hybridization
simulation (8). Hybridization simulation is the computer simulation of a hybridization
of a primer with a specified database. This in silico analysis will identify all hybridization
sites within the database for a candidate primer, allowing the user to select primers
that will be the most specific.
It is important to realize that hybridization simulation is qualitatively different
from a homology or similarity analysis (9). Hybridization simulation uses a thermodynamic
model with nearest neighbor values to calculate the mismatch Tm of hybridization
for all hybridization sites. An example of hybridization simulation data is
shown in Fig. 3.
Currently, hybridization simulation is only available from a single commercially
available program, the HYBsimulator . HYBsimulator allows for the screening of a large
set of candidate primers and selection based on the hybridization simulation data.
3.2.3. Statistical Determination of Specificity
Various mathematical models exist in which the specificity of a given primer can
be estimated based on the frequency of its constituent smaller sequences. One such
method uses a table of frequencies of 6 mers found within a given genomic database