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Fostering new paradigms for the biological sciences

MARK E. BERRES

Genome Center/ Biotechnology Center

Faculty Advisor: Fred Blattner /
Dan Van Der Weide

mark@genome.wisc.edu
262-5578

Resume 2004 (Word Document)

MARK E. BERRES

The use of luminescent quantum dot bioconjugates as

a reporter in microarray assays

      Advances in nonisotopic detection methods have significantly influenced many areas of research ranging from fundamental molecular biology to clinical diagnostics. Most optical methods require fluorescent probes to detect and monitor microscopic interactions. Examples of these labels include many well-known organic fluorophores and naturally occurring chromophores such as green or red fluorescent protein and phycoerytherin. Despite their obvious success in applied research, traditional fluorophores exhibit many undesirable characteristics including relatively weak spectral emissions, short fluorescence lifetimes, narrow excitation and broad emission spectra. Luminescent semiconductor nanocrystals (quantum dots) provide a promising alternative to traditional fluorescent labels. Compared to organic dyes, quantum dots can be orders of magnitude brighter and completely resistant to fading. Dependent only on their physical size, quantum dots also exhibit narrow and symmetrical emission spectra which are excitable over a broad range of wavelengths. These novel optical properties render quantum dots ideal for use in a wide array of biotechnological applications.

      One technology particularly dependent on organic fluorophores as a reporter system is microarrays or "DNA-chips". Microarrays are powerful tools commonly used for high-density genotyping, characterization of gene expression profiles, and more recently, genomic resequencing. However, the sensitivity of conventional microarrays is reduced by poor signal to noise ratios caused, in part, by poor quantum yields and rapid fading of the commonly used fluorophores CY3 and CY5. In terms of the reporter assay itself, biotin end-labeling of nucleic acid, fluorophore-streptavidin bioconjugation, and steric hindrances between DNA and protein molecules further contribute substantial variability to the assay. Each of these phenomena dramatically influences the accuracy and precision within and between microarray experiments. Therefore, a more desirable system would not only include an enhanced reporter but also a simpler assay.

      My primary aim as a GSTP postdoctoral fellow is to incorporate quantum dots into microarray technologies. Specifically, I will develop 1) a means of rapid size-selection of newly synthesized quantum dots, 2) water-soluble surface chemistries compatible with relevant bioconjugation chemistries, and 3) a one-step procedure to deliver bioreactive quantum dots to DNA probes immobilized on a solid substrate. Once completed, I predict that the accuracy and reliability of microarray-based experiments will increase substantially. Advantageously, the use of quantum dots as a reporter assay in microarray experiments will also serve as a model to enhance the performance of other technologies.

 

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