This enables the
PNA probe to bind stronger and the length of the probe can be
shortened. PNA probes have a superior performance compared to DNA probes if regions of the rRNA with complex secondary
structures shall be targeted which have a poor accessibility for
DNA probes PNA probes bind to their
complementary strands even at low salt concentrations and high
temperatures whereas secondary structures consisting out of DNA
or RNA dissolve . Hence, PNA probes can
bind to structures which are hidden from DNA probes. The
accessibility of the rRNA has been shown to possess a great impact
on the fluorescence intensity which might differ significantly
depending on the target region ,and DNA probe
design thus has to take the accessibility into account. The use of
PNA probes (PNA FISH) and suitable hybridization conditions can
circumvent this problem enabling high fluorescence intensities.
The unpolar characteristics of PNA probes also increase the general
penetration properties of the probe, resulting in an enhanced
permeability for the entry into the cell through cell membrane and
cell walls, and, therefore, the hybridization time can be reduced
while keeping high hybridization efficiency. The resistance to
nucleases represents another advantage of PNA probes or other
DNA mimics and RNA derivatives such as locked nucleic acids
(LNA). Accordingly, the use of PNA probes for FISH analysis of food
products has increased significantly in recent years Lately, it was shown for E. coli O157 that PNA probes allow the
specific detection of this particular serotype, which has not been
achieved with a DNA probe The use of
other DNA-mimics (i.e. LNA-DNA hybrid structures with incorporated
LNAs to enhance mismatch discrimination) may promise
similar or even further benefits for FISH-testing in food microbiology
although these mimics have only
rarely been employed. Unfortunately, already published DNA
probes cannot be easily converted into PNA probes or other DNA
mimics since hybridization temperature, buffer stringency and the
number of nucleotides have to be reevaluated and optimized in
each case and self-complementarity has to be avoided. The considerable higher
costs for PNA probes and other DNA-mimics as well as a still
inferior predictability of the results in contrast to conventional
DNA probes also limit their potential for a broader diagnostic use