Modifications of experimental design and parameters, including
shortening of primers and adjustment of PCR conditions, were also
frequently used strategies to increase sensitivity of PCR-based
methods (Dieffenbach, Lowe, & Dveksler, 1993). Proverbially, in
a range of length, shorter primers lead to lower LOD. However,
sensitivity elevation by shortening primers always accompanies
with decrease of method specificity. When the shorter primers
increase binding facility with the target region, the probability of
unexpected non-specific amplification also enhances. In this study,
the sensitivity, specificity and identical efficiency on different
templates could not been simultaneously optimized when the
multiplex PCR primers were truncated. So, shortening primer was
not a feasible approach to lower LOD in this case.