In the present study, we exploited the superior features
of peptide nucleic acids (PNAs) to develop an
efficient PNA zip-code microarray for the detection of
hepatocyte nuclear factor-1a (HNF-1a) mutations that
cause type 3 maturity onset diabetes of the young
(MODY).Amulti-epoxylinkercompoundwassynthesized
and used to achieve an efficient covalent linking
of amine-modified PNA to an aminated glass surface.
PCR was performed to amplify the genomic regions
containing the mutation sites. ThePCRproducts were
then employed as templates in a subsequent multiplex
single base extension reaction using chimeric
primers with 30 complementarity to the specific mutation
site and 50 complementarity to the respective
PNA zip-code sequence on the microarray. The
primers were extended by a single base at each
corresponding mutation site in the presence of
biotin-labeled ddNTPs, and the products were hybridized
to the PNA microarray. Compared to the corresponding
DNA, the PNA zip-code sequence showed a
muchhigher duplex specificity for thecomplementary
DNA sequence. The PNA zip-code microarray was
finally stained with streptavidin-R-phycoerythrin to
generate a fluorescent signal. Using this strategy,
we were able to correctly diagnose several mutation
sites in exon 2 of HNF-1a with a wild-type and mutant
samples including a MODY3 patient. This work represents
one of the few successful applications of
PNA in DNA chip technology.