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PNA Molecular Beacons
DNA is added, the hybridization causes a change in conformation that switches on the
fluorescence since the quencher moves away from the fluorescent unit 19,20 . Depending on the
type of beacon, different strategies are used to keep the probe in the closed form when not
hybridized: DNA beacons are usually kept closed thanks to a stem obtained using two short
complementary sequences at the ends of the probe. PNA molecular beacons may be
maintained closed by an electrostatic interaction exerted by two amino acids bearing opposite
charges at both ends of the probe.
PNA molecular beacons have been used together with many techniques for the recognition of
complementary sequences obtaining a good signal-to-noise ratio, since the flexibility of the
backbone allows a ready closure when no complementary sequence is hybridized. By
coupling these molecules with PNA openers, i.e. PNAs able to strand invade duplex DNA, it
was possible to hybridize not only single stranded DNA, but also double helices making the
target sequence accessible to the PNA. The modified probes display a high selectivity in
sequence recognition, being able to recognize single nucleotide polymorphisms 19 . In order to
increase PNA selectivity, which is necessary in some applications, chiral centers within the
PNA backbone were introduced 21 . A monomer bearing a L-lysine side chain in position 5 has
been introduced in the middle of
N
the sequence, as previous works
demonstrated that enhanced
N
N binding performances are due to
HO O OH
this modification 22 . Moreover,
O
the side chain could be used for
O
HN
O
linking the PNA probe to
O N H O
functional active groups on
O
H
N
PNA
N
NH
H
N
2
surfaces or to nanoparticles 23 .
H
O
The features offered by these
O O
probes allowed their application
NH 3
+
as useful tools in many
Figure 2-4: Chemical structure of a PNA Molecular Beacon (F.
analytical techniques. The
Totsingan, et al., Org. Biomol. Chem., 2008, 6, 1232)
fluorescent signal produced by
the probe after hybridization is very useful in real time PCR, since it allows to monitor the
amplification process in real time 24 . Good performances were obtained also by IE-HPLC 21, 25 ,
by combining the discriminative power offered by the chromatographic technique.
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