Abstract
Peptide nucleic acids (PNAs) are synthetic oligonucleotide analogues based on a pseudopeptide backbone that bind complementary DNA or RNA with high affinity and specificity. In this chapter, three PNA-based genotyping assays are described: PCR clamping, fluorescence-based recognition, and microarray platform. The first two methods are performed in solution, while the microarray method uses a solid surface.
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References
Nielsen PE, Egholm M, Berg RH, Buchardt O (1991) Sequence-selective recognition of DNA by strand displacement with a thymine-substituted polyamide. Science 254:1497–1500
Egholm M, Buchardt O, Christensen L, Behrens C, Freier SM, Driver DA, Berg RH, Kim SK, Norden B, Nielsen PE (1993) PNA hybridizes to complementary oligonucleotides obeying the Watson–Crick hydrogen-bonding rules. Nature 365:566–568
Wittung P, Nielsen PE, Buchardt O, Egholm M, Norden B (1994) DNA like double helix formed by peptide nucleic acid. Nature 368:561–563
Demidov VA, Potaman VN, Frank-KamenetskiJ MD, Egholm M, Buchardt O, Sonnichsen SH, Nielsen PE (1994) Stability of peptide nucleic acids in human serum and cellular extracts. Biochem Pharmacol 48:1310–1313
Sforza S, Corradini R, Tedeschi T, Marchelli R (2011) Food analysis and food authentication by peptide nucleic acid (PNA)-based technologies. Chem Soc Rev 40:221–232
Corradini R, Sforza S, Tedeschi T, Totsingan F, Marchelli R (2007) Peptide nucleic acids with a structurally biased backbone: effects of conformational constraints and stereochemistry. Curr Top Med Chem 7:681–694
Medici MC, Martinelli M, Ruggeri FM, Abelli LA, Bosco S, Arcangeletti MC, Pinardi F, De Conto F, Calderaro A, Chezzi C, Dettori GJ (2005) Broadly reactive nested reverse transcription-PCR using an internal rna standard control for detection of noroviruses in stool samples. J Clin Microbiol 43:3772–3778
Svanvik N, Westman G, Wang D, Kubista M (2000) Light-up probes: thiazole orange-conjugated peptide nucleic acid for detection of target nucleic acid in homogeneous solution. Anal Biochem 281:26–35
Rutjes SA, van den Berg HHJL, Lodder WJ, de Roda Husman AM (2006) Real-time detection of noroviruses in surface water by use of a broadly reactive nucleic acid sequence-based amplification assay. Appl Environ Microbiol 72:5349–5358
Peano C, Lesignoli F, Gulli M, Corradini R, Samson MC, Marchelli R, Marmiroli N (2005) Development of a peptide nucleic acid polymerase chain reaction clamping assay for semiquantitative evaluation of genetically modified organism content in food. Anal Chem 344:174–182
Rozen S, Skaletsky HJ (2000) Primer3 on the WWW for general users and for biologist programmers. In: Krawetz S, Misener S (eds) Bioinformatics methods and protocols: methods in molecular biology. Humana Press, Totowa, NJ, pp 365–386
Kyger EM, Krevolin MD, Powell MJ (1998) Detection of the hereditary hemochromatosis gene mutation by real-time fluorescence polymerase chain reaction and peptide nucleic acid clamping. Anal Biochem 260:142–148
Tedeschi T, Calabretta A, Bencivenni M, Manicardi A, Corrado G, Caramante M, Corradini R, Rao R, Sforza S, Marchelli R (2011) A PNA microarray for tomato genotyping. Mol Biosyst 7:1902–1907
Takiya T, Futo S, Tsuna M, Namimatsu T, Sakano T, Kawai K, Suzuki T (2004) Identification of single base-pair mutation on uidA gene of Escherichia coli O157:H7 by peptide nucleic acids (PNA) mediated PCR clamping. Biosci Biotechnol Biochem 68:360–368
Manicardi A, Calabretta A, Bencivenni M, Tedeschi T, Sforza S, Marchelli R (2010) Affinity and selectivity of C2- and C5-substituted “chiral-box” PNA in solution and on microarrays. Chirality 22:E161–E172
Ørum H, Nielsen PE, Egholm M, Berg RH, Buchardt O, Stanley C (1993) Single base pair mutation analysis by PNA directed PCR clamping. Nucleic Acids Res 21:5332–5336
Tonelli A, Tedeschi T, Germini A, Sforza S, Corradini R, Medici MC, Chezzi C, Marchelli R (2011) Real time RNA transcription monitoring by Thiazole Orange (TO)-conjugated Peptide Nucleic Acid (PNA) probes: norovirus detection. Mol Biosyst 7:1684–1692
Germini A, Rossi S, Zanetti A, Corradini C, Fogher C, Marchelli R (2005) Development of a peptide nucleic acid array platform for the detection of genetically modified organisms in food. J Agric Food Chem 53:3958–3962
Rossi S, Scaravelli E, Germini A, Corradini R, Marchelli R (2006) PNA array platform for the detection of traces of potentially allergenic nuts in foodstuffs. Eur Food Res Technol 223:1–6
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Sforza, S., Tedeschi, T., Bencivenni, M., Tonelli, A., Corradini, R., Marchelli, R. (2014). Use of Peptide Nucleic Acids (PNAs) for Genotyping by Solution and Surface Methods. In: Nielsen, P., Appella, D. (eds) Peptide Nucleic Acids. Methods in Molecular Biology, vol 1050. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-553-8_12
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DOI: https://doi.org/10.1007/978-1-62703-553-8_12
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