DNA sequence detection using surface-enhanced resonance Raman spectroscopy in a homogeneous multiplexed assay.
Detection of specific DNA sequences is central to modern molecular biology and also to molecular diagnostics where identification of a particular disease is based on nucleic acid identification. Many methods exist, and fluorescence spectroscopy dominates the detection technologies employed with different assay formats. This study demonstrates the use of surface-enhanced resonance Raman scattering (SERRS) to detect specific DNA sequences when coupled with modified SERRS-active probes that have been designed to modify the affinity of double- and single-stranded DNA for the surface of silver nanoparticles resulting in discernible differences in the SERRS which can be correlated to the specific DNA hybridization event. The principle of the assay lies on the lack of affinity of double-stranded DNA for silver nanoparticle surfaces; therefore, hybridization of the probe to the target results in a reduction in the SERRS signal. Use of locked nucleic acid (LNA) residues in the DNA probes resulted in greater discrimination between exact match and mismatches when used in comparison to unmodified labeled DNA probes. Polymerase chain reaction (PCR) products were detected using this methodology, and ultimately a multiplex detection of sequences relating to a hospital-acquired infection, namely, methicillin-resistant Staphylococcus aureus (MRSA), demonstrated the versatility and applicability of this approach to real-life situations.
MacAskill
Alexandra
A
Centre for Molecular Nanometrolgy, WestCHEM, Pure and Applied Chemistry, University of Strathclyde, 295 Cathedral Street, Glasgow, G1 1XL, UK.
Crawford
David
D
Graham
Duncan
D
Faulds
Karen
K
D001483:Base Sequence; D004247:DNA / Q000032:analysis* / Q000737:chemistry; D015342:DNA Probes / Q000737:chemistry; D053768:Metal Nanoparticles / Q000737:chemistry; D055624:Methicillin-Resistant Staphylococcus aureus / Q000235:genetics / Q000302:isolation & purification; D009693:Nucleic Acid Hybridization / Q000379:methods; D009841:Oligonucleotides / Q000737:chemistry; D012834:Silver / Q000737:chemistry; D013059:Spectrum Analysis, Raman / Q000379:methods*
D015342:DNA Probes; D009841:Oligonucleotides; C477371:locked nucleic acid; D012834:Silver; D004247:DNA