Cornell University

The University of North Carolina at Chapel Hill

Louisiana State University and Agricultural & Mechanical College

John William Efcavitch

Francis Barany

Steven A Soper

Sunggook Park

Michael C Murphy

Jianmin Huang

Collin J McKinney

Mateusz Hupert

Philip B Feinberg

Cristian Ruiz Rueda

Michael Murphy

The present invention relates to a method for the highly specific, targeted capture of regions of human genomes and transcriptomes from the blood, i.e. from cell free circulating DNA, exosomes, microRNA, circulating tumor cells, or total blood cells, to allow for the highly sensitive detection of mutation, expression, copy number, translocation, alternative splicing, and methylation changes using combined nuclease, ligation, polymerase, and massively parallel sequencing reactions. The method generates a collection of different circular chimeric single-stranded nucleic acid constructs, suitable for sequencing on multiple platforms. In some embodiments, each construct of the collection comprised a first single stranded segment of original genomic DNA from a host organism and a second single stranded synthetic nucleic acid segment that is linked to the first single stranded segment and comprises a nucleotide sequence that is exogenous to the host organism. These chimeric constructs are suitable for identifying and enumerating mutations, copy changes, translocations, and methylation changes. In other embodiments, input mRNA, lncRNA, or miRNA is used to generate circular DNA products that reflect the presence and copy number of specific mRNA's, lncRNA's splice-site variants, translocations, and miRNA.

Method for identification and enumeration of nucleic acid sequence, expression, copy, or DNA methylation changes, using combined nuclease, ligase, polymerase, and sequencing reactions

The present invention relates to methods and devices for identifying and quantifying, including low abundance, nucleotide base mutations, insertions, deletions, translocations, splice variants, miRNA variants, alternative transcripts, alternative start sites, alternative coding sequences, alternative non-coding sequences, alternative splicings, exon insertions, exon deletions, intron insertions, or other rearrangement at the genome level and/or methylated nucleotide bases.

Method for identification and quantification of nucleic acid expression, splice variant, translocation, copy number, or methylation changes

The present invention is directed methods for identifying, in a sample, one or more target nucleotide sequences differing from other nucleotide sequences in the sample by one or more nucleotides, one or more copy numbers, one or more transcript sequences, and/or one or more methylated residues, using ligation detection reactions, polymerase mediated extension reactions, and/or cleavage reactions. The present invention is also directed to methods for identifying, in a sample, one or more nucleotides in a target nucleotide sequence.

Method for identification and enumeration of nucleic acid sequences, expression, splice variant, translocation, copy, or DNA methylation changes using combined nuclease, ligase, polymerase, terminal transferase, and sequencing reactions

The present invention is directed to methods comprising a device that comprises a biomolecular processor and one or more nanotubes. Each biomolecular processor comprises a bioreactor chamber defined by a solid substrate, a plurality of spaced support structures within said bioreactor chamber and attached to the solid substrate, one or more nanotubes defined by the solid substrate and fluidically coupled to the bioreactor chamber and one or more capture molecules immobilized to some or all of said plurality of spaced support structures, said one or more capture molecules suitable to bind to a portion of a target nucleic acid molecule in a sample. The nanotubes have a passage extending between an input end proximate to the bioreactor chamber and an output end distal to the bioreactor chamber, and comprises one or more nanopores within the passage with each nanopore having a reduced diameter relative to the passage.

Universal molecular processor for precision medicine

The present invention relates to a method for the highly specific, targeted capture of regions of human genomes and transcriptomes from the blood, i.e. from cell free circulating DNA, exosomes, microRNA, circulating tumor cells, or total blood cells, to allow for the highly sensitive detection of mutation, expression, copy number, translocation, alternative splicing, and methylation changes using combined nuclease, ligation, polymerase, and massively parallel sequencing reactions. The method generates a collection of different circular chimeric single-stranded nucleic acid constructs, suitable for sequencing on multiple platforms. In some embodiments, each construct of the collection comprised a first single stranded segment of original genomic DNA from a host organism and a second single stranded synthetic nucleic acid segment that is linked to the first single stranded segment and comprises a nucleotide sequence that is exogenous to the host organism. These chimeric constructs are suitable for identifying and enumerating mutations, copy changes, translocations, and methylation changes. In other embodiments, input mRNA, IncRNA, or miRNA is used to generate circular DNA products that reflect the presence and copy number of specific mRNA's, IncRNA's splice-site variants, translocations, and miRNA.

The present invention is directed to a device that comprises a biomolecular processor and one or more nanotubes. Each biomolecular processor comprises a bioreactor chamber defined by a solid substrate, a plurality of spaced support structures within said bioreactor chamber and attached to the solid substrate, and one or more capture molecules immobilized to some or all of said plurality of spaced support structures, said one or more capture molecules suitable to bind to a portion of a target nucleic acid molecule in a sample. The device also comprises one or more nanotubes defined by the solid substrate and fluidically coupled to the bioreactor chamber.

Growing community of inventors

San Carlos, CA, United States of America

John William Efcavitch