Applied Biosystems, LLC

Xtrana, Inc.

Applera Corporation

Molecular Innovations, Inc.

Other

Visugen Global LLC

Life Technologies Corporation

John C Gerdes

Jeffrey M Marmaro

Elaine A Best

Jeffery M Marmaro

Christopher A Roehl

Jeffrey T Ives

Diane L Kozwich

Kris S Buchanan

Kirsten L Nelson

Lynn D Jankovsky

Systems and methods that enable the rapid identification of target molecules present in a sample at low concentrations is provided. The system includes a sample volume, and a detection structure that is connected to the sample volume by a conduit. The detection structure includes a microfluidic chip that defines a plurality of fluid channels. The walls of the fluid channels are formed from or covered with a metal oxide to which target molecules attach. After the sample volume has been passed through the detection structure, and in particular through the channels, visible microparticles are passed through the detection structure. The visible microparticles are configured to have an affinity for the target molecules. The microfluidic chip is then imaged to detect visible microparticles, and to thereby obtain information regarding the presence of the target molecules.

Systems and methods for capture and detection of low copy targets from large sample volumes

A two-step multiplex amplification reaction includes a first step which truncates the standard initial multiplex amplification round to 'boost' the sample copy number by only a 100-1000 fold increase in the target. Following the first step the product is divided into optimized secondary single amplification reactions, each containing one of the primer sets that were used previously in the first or multiplexed booster step. The booster step can occur using an aqueous target nucleic acid or using a solid phase archived nucleic acid. In particular, nucleic acid sequences that uniquely identifywere identified using the multiplex amplification method.

Methods for multiplexing amplification reactions

The present invention provides a two-step multiplex amplification reaction wherein the first step truncates the standard initial multiplex amplification round to 'boost' the sample copy number by only a 100-1000 fold increase in the target. Following the first step the product is divided into optimized secondary single amplification reactions, each containing one of the primer sets that were used previously in the first or multiplexed booster step. The booster step can occur using an aqueous target nucleic acid or using a solid phase archived nucleic acid. In particular, nucleic acid sequences that uniquely identifywere identified using the multiplex amplification method.

Methods and devices for multiplexing amplification reactions

This invention is directed to a process for tightly binding nucleic acid to solid phase and corresponding processes for the utilization thereof. Nucleic acid is bound to solid phase matrices exhibiting sufficient hydrophilicity and electropositivity to tightly bind the nucleic acids from a sample. These processes include nucleic acid (double or single stranded DNA and RNA) capture from high volume and/or low concentration specimens, buffer changes, washes, and volume reductions, and enable the interface of solid phase bound nucleic acid with enzyme, hybridization or amplification strategies. The tightly bound nucleic acid may be used, for example, in repeated analyses to confirm results or test additional genes in both research and commercial applications. Further, a method is described for virus extraction, purification, and solid phase amplification from large volume plasma specimens.

Nucleic acid archiving

The present invention provides a two-step multiplex amplification reaction wherein the first step truncates the standard initial multiplex amplification round to 'boost' the sample copy number by only a 100&#x2013;1000 fold increase in the target. Following the first step the product is divided into optimized secondary single amplification reactions, each containing one of the primer sets that were used previously in the first or multiplexed booster step. The booster step can occur using an aqueous target nucleic acid or using a solid phase archived nucleic acid. In particular, nucleic acid sequences that uniquely identifywere identified using the multiplex amplification method.

This invention provides a kit comprising a substrate having a surface coated with a solid phase matrix for nucleic acid manipulation. The solid phase matrix exhibits sufficient hydrophilicity and electropositivity to tightly bind the nucleic acids in a sample. The manipulations include nucleic acid (double or single stranded DNA and RNA) capture from high volume and/or low concentration specimens, buffer changes, washes, and volume reductions, and enable the interface of solid phase bound nucleic acid with enzyme, hybridization or amplification strategies. The tightly bound nucleic acid may be used, for example, in repeated analyses to confirm results or test additional genes in both research and commercial applications. Further, a method for virus extraction, purification, and solid phase amplification from large volume plasma specimens is described.

Self-contained devices are described that integrate nucleic acid extraction, specific target amplification and detection into a single device. This integration permits rapid and accurate nucleic acid sequence detection. The invention may be used, for example, in the screening for nucleic acid sequences which may be indicative of genetic defects or contagious diseases, as well as for monitoring efficacy in the treatment of contagious diseases.

Self-contained device integrating nucleic acid extraction, amplification and detection

The present invention pertains to methods, reagents, compositions, kits, and instruments for use in simultaneously amplifying multiple targets. In particular, this invention is based on the discovery of a two-step multiplex amplification reaction wherein the first step truncates the standard initial multiplex amplification round to &ldquo;boost&rdquo; the sample copy number by only a 100-1000 fold increase in the target. Following the first step the product is divided into optimized secondary single amplification reactions, each containing one of the primer sets that were used previously in the first or multiplexed booster step. The booster step can occur using an aqueous target nucleic acid or using a solid phase archived nucleic acid.

This invention is directed to a process for irreversibly binding nucleic acid to solid phase and corresponding processes for the utilization thereof. Nucleic acid is bound to solid phase matrixes exhibiting sufficient hydrophilicity and electropositivity to irreversibly bind the nucleic acids from a sample. These processes include nucleic acid (double or single stranded DNA and RNA) capture from high volume:low concentration specimens, buffer changes, washes, and volume reductions, and enable the interface of solid phase bound nucleic acid with enzyme, hybridization or amplification strategies. The invention, solid phase irreversibly bound nucleic acid, may be used, for example, in repeated analyses to confirm results or test additional genes in both research and commercial applications. Further, a method is described for virus extraction, purification, and solid phase amplification from large volume plasma specimens.

This invention is directed to methods for the quantitative measurement of specific gene expression levels in biological samples. In one embodiment, methods for the quantitative monitoring of gene expression without either co-amplification of an added template or use of an endogenous constitutive transcript are provided. The former involves a duplex amplification reaction in which a single set of primers is used to amplify both genomic DNA and expressed mRNA from the same gene sequence. These primers are targeted for sequences flanking the splice junction and intron sequences for the mRNA and DNA respectively. By their use, any suitable nucleic acid amplification technology yields mRNA and DNA amplimers which are distinguishable by length and sequence heterogeneity. These amplimers are also present in the final amplification reaction in ratios which are dependent upon the ratios of the expressed mRNA to the DNA in the sample, allowing the quantitation of mRNA in a sample which is normalized to the number of copies of genomic DNA since the genomic DNA acts as the internal quantitation standard, and in effect yields the amount of mRNA per cell. Any detection methodology which can detect amplimers of different lengths or sequences can be used for post amplification quantitation. This strategy may be employed for any gene system in which the mRNA sequence differs from the original genomic DNA sequence. In another embodiment, methods for the quantitative monitoring of gene expression involving determining the ratio of genomic material to expressed mRNA without nucleic acid amplification and/or primer binding are described. This method includes the independent and direct detection of the genomic material and mRNA by complementary probe binding without prior amplification. Additional methods for quantitative measurement of gene expression rates without RNA transcription region introns are described. The invention is useful in gene expression determination in research and commercial applications.

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Denver, CO, United States of America

John C Gerdes