Nugen Technologies, Inc.

Syntex (u.s.a.) Inc.

Behringwerke Aktiengesellschaft

Dade Behring Inc.

Dade Behring Marburg Gmbh

Aventis Behring Gmbh

Behring Diagnostics Gmbh

Siemens Healthcare Diagnostics Products Gmbh

Tecan Genomics, Inc.

Nurith Kurn

Edwin F Ullman

Rajesh D Patel

Martin J Becker

Shenglong Wang

Arthur C Switchenko

Alla Lishanski

Marcel R Pirio

Doug Amorese

Donald E Berger, Jr

Yen Ping Liu

Christian Neukom

Vartan E Ghazarossian

Litai Weng

Chris Armour

N Chung Sizto

Geoffrey A Dafforn

Sharat Singh

Dariush Davalian

Bin Li

Samuel J Rose

Cheng-I Lin

John S Pease

Marc Taylor

John D Olson

Christopher D Armour

Remy Cromer

Thomas M Houts

Steve De Keczer

Harshvardhan B Mehta

Chander Bahl

Pengchin Chen

Martin Junhong Wang

Claire Lin

Rajesh Patel

The present invention provides methods, compositions and kits for targeted nucleic acid sequence enrichment in a nucleic acid sample and for high efficiency nucleic acid library generation for next generation sequencing (NGS). Specifically, the methods, compositions and kits provided herein are useful for the production and capture of amplification-ready, target-specific and strand-specific regions of interest from nucleic acid samples containing complex DNA.

Compositions and methods for targeted nucleic acid sequence enrichment and high efficiency library generation

The present invention provides methods, compositions and kits for the generation of next generation sequencing (NGS) libraries in which non-desired nucleic acid sequences have been depleted or substantially reduced. The methods, compositions and kits provided herein are useful, for example, for the production of libraries from total RNA with reduced ribosomal RNA and for the reduction of common mRNA species in expression profiling from mixed samples where the mRNAs of interest are present at low levels. The methods of the invention can be employed for the elimination of non-desired nucleic acid sequences in a sequence-specific manner, and consequently, for the enrichment of nucleic acid sequences of interest in a nucleic acid library.

Compositions and methods for negative selection of non-desired nucleic acid sequences

Compositions and methods for targeted nucleic acid sequence enrichment and high efficiency library regeneration

The invention provides methods for isothermal amplification of RNA. The methods are particularly suitable for amplifying a plurality of RNA species in a sample. The methods employ a composite primer, a second primer and strand displacement to generate multiple copies of DNA products comprising sequences complementary to an RNA sequence of interest. In another aspect, the methods employ a single primer (which is a composite primer) and strand displacement to generate multiple copies of DNA products comprising sequences complementary to an RNA sequence of interest. In some embodiments, a transcription step is included to generate multiple copies of sense RNA of an RNA sequence of interest. The methods are useful for preparation of nucleic acid libraries and substrates for analysis of gene expression of cells in biological samples. The invention also provides compositions and kits for practicing the amplification methods, as well as methods which use the amplification products.

Compositions for amplification of RNA sequences using composite primers

The invention relates to the field of polynucleotide amplification. More particularly, the invention provides methods, compositions and kits for amplification of (i.e., making multiple copies of) a multiplicity of different polynucleotide template sequences using a randomly primed RNA/DNA composite primer.

Global amplification using a randomly primed composite primer

The invention provides methods for amplification of polynucleotide sequences using primers containing single-stranded RNA. The methods employ use of an enzyme capable of cleaving single-stranded RNA, such as RNase I, to degrade a first RNA-containing primer prior to addition of a second RNA-containing primer. The invention also provides compositions and kits for practicing the amplification methods, as well as methods which use the amplification products.

Nucleic acid amplification procedure using RNA and DNA composite primers

The invention provides methods, compositions, and kits for fragmentation and labeling of nucleic acids. More particularly, the invention relates to methods for fragmentation of nucleic acids to produce fragments with 3' end hydroxyl groups within a desired size range. In methods of the invention, nucleic acids are fragmented at abasic sites to produce fragments with blocked 3&#x2032; ends. The 3&#x2032; ends are unblocked to produce polynucleotide fragments with hydroxyl groups at their 3&#x2032; ends. Methods, kits, and compositions for carrying out fragmentation of a polynucleotide template in a single reaction mixture to yield fragments with 3&#x2032;-hydroxyl ends within the desired size range are disclosed.

Methods for fragmentation and labeling of nucleic acids

The present method provides methods, libraries, and kits related to the archiving and clonal amplification of sequences related to target polynucleotide sequences. The method allows for the generation and attachment of polynucleotides with defined 3' and 5&#x2032; ends to solid surfaces. The polynucleotides attached to the solid substrates can be stored or archived as libraries and can subsequently be retrieved for analysis, for example by clonal amplification using a single composite amplification primer comprising a DNA portion and an RNA portion. In some embodiments, nucleotides attached to solid surfaces can be used for sequencing of nucleotide sequences related to the target DNA. The methods are applicable to total RNA and/or total DNA analysis.

Method for clonal amplification

Methods and compositions for amplification of RNA sequences

The present invention provides novel isothermal methods of generating multiple copies of, detecting and/or quantifying nucleic acid sequences of interest based on limited primer extension or attachment of oligonucleotide pairs using composite RNA/DNA primers. Methods for generating multiple copies of and/or detecting and/or quantifying nucleic acid sequences, wherein products of primer extension or attachment of oligonucleotide pairs comprising a cleavable portion are generated, and wherein cleavage of the products results in dissociation of cleaved products from target polynucleotides, are provided. The invention further provides compositions, kits and systems for practicing these methods.

Methods and compositions for generation of multiple copies of nucleic acid sequences and methods of detection thereof

The invention relates to methods for analysis of nucleic acid methylation status, and fragmentation and/or labeling and/or immobilization of nucleic acids. More particularly, the invention relates to methods for fragmentation and/or labeling and/or immobilization of nucleic acids comprising labeling and/or cleavage and/or immobilization at abasic sites.

Methods for analysis of nucleic acid methylation status and methods for fragmentation, labeling and immobilization of nucleic acids

Methods and compositions are provided related to the amplification of target polynucleotide sequences as well as total RNA and total DNA amplification. In some embodiments, the methods and compositions also allow for the immobilization and capture of target polynucleotides with defined 3' and or 5&#x2032; sequences to solid surfaces. The polynucleotides attached to the solid surfaces can be amplified or eluted for downstream processing. In some cases, nucleotides attached to solid surfaces can be used for high throughput sequencing of nucleotide sequences related to target DNA or target RNA.

Isothermal nucleic acid amplification methods and compositions

Growing community of inventors

Palo Alto, CA, United States of America

Nurith Kurn