View - DSpace UniPR

Arg-PNA Microarrays
4.2 Results and Discussion
4.2.1 Design and synthesis of modified chiral PNA probes
The properties of modified chiral PNAs containing lysine based monomers in the middle of
the sequence have been showed in chapter 3, demonstrating that PNAs containing one residue
with two lysine-derived side chains (2D,5L) had a highly improved affinity and selectivity in
the recognition of complementary DNA sequences. An homologous PNA having the same
modification but based on the structure of arginine maintained the same properties.
As explained in chapter 3, arginine has been used instead of lysine because the PNAs will be
linked to surface for building PNA-microarrays. The presence of the guanidium group on the
side chain of arginine allows to avoid any interference in the reaction between the terminal
amino group of the probe and the reactive groups on the surface (which was possible if using
lysine-based PNAs).
The PNA sequences to be synthesized had to be complementary to the two gene tracts bearing
the SNPs, codifying for the three ApoE protein isoforms. Four undecameric PNAs (for two
different SNPs in two different gene tracts) were designed (PNA 1-4, Table 4-1), with the
chiral (2D,5L-Arg) unit located in the middle of the sequence, corresponding to the SNP
positions. In both regions containing the codons for amino acid 112 and 158 in the ApoE
protein, a T/C SNP was present, thus in both cases the PNA nucleobase corresponding to the
SNP was an A or a G. It was also decided to place two spacer groups
(aminoethoxyethoxyacetyl, AEEA) at the amino terminus in agreement with previous
studies 27 which established the right distance between the probes and the surface of
microarray for obtaining a good surface hybridization.
77