SNPs at a snip: SERS spots singles

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  • Published: Aug 1, 2011
  • Author: David Bradley
  • Channels: Raman
thumbnail image: SNPs at a snip: SERS spots singles

Picking up good variations

Surface enhanced Raman spectroscopy can now be used to detect single nucleotide substitutions (the all-important SNPs of genetic variance and personalised medicine) in DNA without the need for labels or indeed any chemical modification of the oligonucleotides.

Evanthia Papadopoulou and Steven Bell of the Innovative Molecular Materials Group, in the School of Chemistry & Chemical Engineering at Queen's University Belfast, UK, explain how DNA sequences can be induced to spontaneously adsorb to the surfaces of silver colloids through their nucleotide side chains. This then facilitates surface-enhanced Raman spectroscopy (SERS) revealing details of these non-specifically bound strands. The modification is sufficiently reproducible that the technique can be used to identify single-base mismatches in short (25-mer and 23-mer) strands, the researchers say.

A single-nucleotide polymorphism is a variation in the DNA sequence that occurs when one base - G, A, T, C - is different between members of a biological species or paired chromosomes in an individual. For example, there is a SNP between the short DNA sequences: AAGCCTA and AAGCTTA.

SNP associations

There are SNPs associated with a tendency to have a particular physical trait such as baldness, athleticisim, green eyes, red hair, obesity, alcohol dependence, type 2 diabetes and many other diseases, even the amount of earwax one produces. There are also SNPs associated with the presence of metabolic enzymes or the lack thereof that mean an individual is more or less responsive to a given pharmaceutical or other therapeutic agent. There are often variations between human populations, so a SNP that is common in one geographical or ethnic group may be much rarer in another.

"Detection of SNPs is already fundamental to diagnosis of genetic disease," Bell told SpectroscopyNOW, "For example, more than half of all known disease mutations come from replacement polymorphisms."

Bell adds that, "SNPs can affect an individual's susceptibility to common diseases. This means that in the future more powerful analytical tools could potentially allow personalised healthcare where a person's susceptibility to various diseases might be determined by the familly doctor in their surgery, allowing preventitive measure to be put in place." Moreover, even before that is possible, there is still the opportunity for introducing bedside diagnostics into hospitals. The potential is thus driving the search for cost-effective approaches to DNA analysis.

"MALDI-TOF" mass spectrometry (matrix-assisted laser desorption/ionization time-of-flight MS) is currently used widely to detect SNPs because it is label free and highly sensitive. However, it is also highly expensive. Bell and colleagues reasoned that the much less costly SERS might be an appropriate alternative that would also be highly sensitive and label free.

Previously, Bell's team had developed the technology to carry out direct label-free identification of mononucleotides in aqueous solution using SERS; the SERS signals are very large for the negatively charged colloidal particles aggregated with magnesium sulfate. "There is no requirement for labels because the Raman signals of each of the mononucleotides are intrinsically different due to the differences in their chemical structures, Bell explained at the time, adding that the novel strategy could be useful in sequencing strands of DNA.

Going label free

"We can now obtain the difference spectra for DNA sequences containing features corresponding to the exchanged nucleotides," Bell explains. This means that SERS could be used to identify the same differences in natural DNA sequences, in others words to identify SNPs. "The best way to highlight the changes in the spectra is to digitally subtract them, which should remove the contributions from unchanged nucleotides to give difference spectra that contain positive and negative features corresponding to the exchanged nucleotides," the team explains. After all, each polymorphism gives a different SERS band and even small changes can be revealed in this way.

"Of course our work is still very much basic research and taking advantage of these observations will require the development of a whole technology," concedes Bell. "But, it is worth remembering that when we started on this path a few years ago the typical large size (large benchtop) and high cost (ca. £100k) of the instruments needed to read the signals was prohibitive." He and his colleagues realised that was likely to change and now his team uses hand-held Raman spectrometers in the lab that weigh less than 500 g and cost a mere £10k. The next generation of devices will be smaller still and cheaper.

"Against this background it makes sense to continue pushing forward the chemistry," Bell adds. "There is already very elegant work being carried out using SERS to detect specific DNA sequences using labelled materials, such as that from Duncan Graham's group in Strathclyde." Bell hopes that his team's work can complement such efforts by going for appplications where the low cost and high speed of label-free methods can make them the method of choice. "As a spectroscopist at heart I just like the simplicity of detecting the bases which are present by reading their signals directly in the surface-enhanced Raman spectra," he says.

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

Surface enhanced Raman spectroscopy can now be used to detect single nucleotide substitutions (the all-important SNPs of genetic variance and personalised medicine) in DNA without the need for labels or indeed any chemical modification of the oligonucleotides.
Green eyes and red hair?
Blame your SNPs

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