Diagnostic Biological Tests/Assays/Genomics/Pretomics

Dr. Smith has developed a new class of water-stable nanoplatelets encapsulated in lipoproteins. Nanoplatelets are bright single-molecule emitters, comparable to quantum dots, but with substantially narrower bandwidths. The nanoparticles retain their fluorescence after internalization in cells and thus are suitable for live-cell single-molecule imaging.

This invention is a novel approach to performing DNA sequencing-by-synthesis methodology that is anticipated to make DNA sequencing routine, highly accurate and affordable.

The approach is a silicon-based field effect nanoscale sensor array using a sequence-by-synthesis methodology. PCR is performed locally in each pixel through the use of recently developed methods for heating fluid using a silicon field effect sensor, and then the amplified molecules in the pixel are detected in a label-free electrical manner where the added charge during the synthesis process is detected using the same thin-film SOI transistor.

Cost is significantly reduced since the nucleotides do not need a fluorescent label, there is no need for gel electrophoresis, and the entire machinery of optical signal processing is unnecessary. In addition, since the silicon field effect sensors when used in a nanoscale dimension can be very sensitive and detect the charge from just a few molecules, it is quite feasible that the PCR amplification can be utilized with very few cycles. Reduction of PCR cycling would reduce the usage of costly reagents. The information retrieval can be scaled and the information obtained per unit time is maximized.

Researchers from the University of Illinois have identified a new, cheaper and easier way to detect methylation (alterations) of DNA. By measuring the methylation of certain DNA sequences, doctors can predict the likelihood of cancers and other disorders. This early detection has the potential to save lives. 

Traditional methods for forming carbon-carbon bonds between unsaturated moieties have a number of well-known difficulties. These cross-coupling reactions often require harsh conditions, use difficult-to-handle precursors, and result in poisonous byproducts. This invention uses known materials-organosilacyclobutanes or simple organosilanols-in a new way to avoid these problems. The process involves subjecting these materials to stereospecific cross-coupling reactions with organic electrophiles in the presence of an activator (either a fluoride source or oxygen-based nucleophile) and a palladium catalyst. Under these mild conditions, carbon-carbon bonds are formed without the toxic byproducts associated with traditional cross-coupling methods.

This invention provides a mild, general and stereospecific method for the formation of carbon-carbon bonds between unsaturated moieties without the concomitant production of toxic byproducts. This new method addresses many of the difficulties associated with traditional cross-coupling techniques. By using organosilacyclobutanes or organosilanols, this process eliminates the need for processing in harsh conditions and using difficult-to-handle precursors. Most importantly, it eliminates the toxic byproducts that result from traditional methods. The cross-coupling begins with an organosilacyclobutane or organosilanol that is treated with a fluoride solution. After the initial exotherm subsides, the organic electrophile is added along with the palladium catalyst. The reactions times vary from 10 minutes to 7 hours at room temperature. The products then are isolated by removal of solvent and filtration through silica gel to remove salts and catalyst, followed by distillation. Occasionally filtration through a reverse phase (C-18) chromatography column is necessary to remove the byproduct. The products are formed in 65% to 93% of analytically pure material.

Organosilacyclobutanes or organosilanols are a preferable material for cross-coupling reactions:

• Unlike magnesium and lithium agents, these materials are chemically inert and offer good functional-group compatibilities.
• Unlike zinc and aluminum agents, these materials are easy to prepare and generate in a geometrically defined form.
• Unlike tin agents, these materials do not result in toxic, heavy-metal byproducts.
• Unlike boron agents, these materials are not air sensitive and can be readily carried through synthetic sequences.
• Unlike previously employed organosilicon reagents, these materials have good reactivity.
• Unlike fluorosilanes, these materials can be processed at room temperature and are based on easy-to-handle precursors. Unlike orthosiliconates, these materials are not water sensitive and are highly reactive.

Benefits

The benefits of this invention accrue from its use of organosilacyclobutanes or organosilanols as the nucleophilic component in cross-coupling reactions. Unlike the materials used in traditional processes, these silicon-based moieties are chemically inert, have low molecular weight, are easy to introduce in a variety of organic structures, are stable to standard purification procedures, and can be activated by fluoride ions at room temperature. These features make the coupling process less complicated and less dangerous than traditional coupling methods.

• Simple: This invention eliminates the need for high-temperature processing conditions. Also, the silicon-based materials are easy to synthesize and purify.
• Low risk: This new process does not require the use of difficult-to-handle precursors traditionally needed for the formation of carbon-carbon bonds. Organosilacyclobutanes and organosilanols are chemically inert, while the fluoride source and palladium catalyst offer mild reaction conditions.
• Nontoxic: Unlike tin agents, silicon-based materials have a low molecular weight and do no yield heavy-metal byproducts. Thus, this process can be used in large-scale applications without concern for product contamination or toxic waste streams.
• Robust: Unlike boron agents, silicon-based materials are not air sensitive and can be carried through synthetic sequences readily.