Label-free selective DNA detection with high mismatch recognition by PNA beacons and ion exchange HPLC.
Two 11mer peptide nucleic acid (PNA) beacons were synthesized and tested for the detection of full-matched or single mismatched DNA. Fluorescent measurements carried out in solution showed only partial discrimination of the mismatched sequence, while using anion-exchange HPLC, in combination with fluorimetric detection, allowed DNA analysis to be performed with high sensitivity and extremely high sequence selectivity. Up to >90 : 1 signal discrimination in the presence of one single mismatched base was observed. The analysis was tested on both short and long DNA oligomers. Detection of DNA obtained from PCR amplification was also performed allowing the selective detection of the target sequence in complex mixtures. Label free detection of the DNA with high sequence selectivity is therefore possible using the present approach.
Totsingan
Filbert
F
Dipartimento di Chimica Organica e Industriale-Università di Parma, Viale G.P. Usberti 17/a-I43100, Parma, Italy.
Rossi
Stefano
S
Corradini
Roberto
R
Tedeschi
Tullia
T
Sforza
Stefano
S
Juris
Alberto
A
Scaravelli
Elena
E
Marchelli
Rosangela
R
D020137:Base Pair Mismatch; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000032:analysis / Q000737:chemistry*; D007475:Ion Exchange Resins / Q000737:chemistry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D020135:Peptide Nucleic Acids / Q000737:chemistry*
D007475:Ion Exchange Resins; D020135:Peptide Nucleic Acids; D004247:DNA