Application of a lanthanide fluorescent chelate label for detection of single-nucleotide mutations with peptide nucleic acid probes.
We developed the approach to detect single-nucleotide mutation with peptide nucleic acid (PNA) probes and time-resolved fluorometry using a fluorescence lanthanide chelate label, {2,2',2'',2'''-{4'-{[(4,6-dichloro-1,3,5-triazin-2-yl)amino]biphenyl-4-yl}-2,2': 6',2''-terpyridine-6,6''-diyl}bis(methylenenitrilo)}tetrakis(acetato)}europium(III) (DTBTA-Eu3+). Compared with DNA probes, PNA probes showed lower mismatch signals and gave higher signal/noise (S/N) ratios. Using the system, we examined the single-nucleotide mutations of codon 12 in the c-Ha-ras gene of PCR amplicons of genome DNAs isolated from human umbilical vein endothelial cells (HUVECs) and T24 cells.
Hashino
Kimikazu
K
Advanced Research Institute for Science and Engineering, Waseda University, Okubo, Shinjuku-ku, Tokyo 169-8555, Japan.
Ito
Masahiro
M
Ikawa
Keisuke
K
Hosoya
Chihiro
C
Nishioka
Takuya
T
Matsumoto
Kazuko
K
D001483:Base Sequence; D002614:Chelating Agents / Q000737:chemistry*; D003062:Codon / Q000235:genetics; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry*; D005470:Fluorometry / Q000379:methods; D005784:Gene Amplification / Q000235:genetics; D011905:Genes, ras / Q000235:genetics; D016678:Genome / Q000235:genetics; D006801:Humans; D028581:Lanthanoid Series Elements / Q000737:chemistry*; D008969:Molecular Sequence Data; D009154:Mutation; D015341:Nucleic Acid Probes / Q000032:analysis* / Q000235:genetics; D009711:Nucleotides / Q000032:analysis*; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D014471:Umbilical Veins / Q000166:cytology
D002614:Chelating Agents; D003062:Codon; D005456:Fluorescent Dyes; D028581:Lanthanoid Series Elements; D015341:Nucleic Acid Probes; D009711:Nucleotides