Gold-Coupled Nucleotides for Ultrasensitive Probes
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25 East Loop Road, Suite 124, Stony Brook, NY, 11790
Richard D. Powell
AbstractNew nucleotide reagents are proposed that have covalently attached 0.8 or 1.4 nm gold clusters. These will then be used to develop extremely sensitive and high resolution hybridization probes. Several synthetic pathways will be explored to produce gold labelled uridine and adenine nucleotide triphosphates. Phophoramidite - gold derivatives will also be synthesized. The first set of labels will be incorporated enzymatically into nucleic acids to create gold-labelled hybridization probes. The phosphoramidite - gold labels will be utilized in automatic DNA synthesizers for the synthesis of labelled oligonucleotides. The efficiency of gold- nucleotide incorporation will be determined, and the most efficient probes will be employed in dot blots and in situ hybridization tests to localize nucleic acids in mammalian cells at high resolution. Double labelling with the 1.4 nm and 0.82 nm probes will enable the localization of two different nucleic: add sequences within the same cell. Gold labelled probes will enhance the sensitivity an order of magnitude or more over current probes. Improved penetration up to 30 micromoles into cells and tissue sections will also be realized with the new probes. Applications of this new technology should improve genetic topography studies in light and electron microscopy, Southern blotting, gene mapping and sequencing, and clinical diagnosis.
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