Abstract: The present invention relates to methods of introducing multiple expression constructs into a eukaryotic cell, methods of constructing a eukaryotic cell having multiple target loci for expressing multiple heterologous proteins of interest, eukaryotic cells for expressing multiple heterologous proteins of interest, and methods of production of multiple heterologous proteins of interest.
Type:
Application
Filed:
March 9, 2016
Publication date:
February 8, 2018
Applicant:
Novozymes A/S
Inventors:
Qiming Jin, Jeffrey Shasky, Donna Moyer, Abigail Jang, Gloria Muzzi-Erichsen, Cara Kleindienst
Abstract: Methods and compositions are provided for displaying a protein of interest (POI) on the surface of a eukaryotic cell by fusing the POI to a signal polypeptide, a stalk polypeptide, and a surface anchor polypeptide to generate a surface accessible fusion protein. Nucleic acids are provided that include nucleotide sequences encoding a signal polypeptide, a stalk polypeptide, and a surface anchor polypeptide. In some cases, a subject nucleic acid includes and insertion site for the insertion of a POI. In some cases, a subject nucleic acid includes a nucleotide sequence that encodes a POI. In some cases a stalk polypeptide is a synthetic stalk polypeptide and various example synthetic stalk polypeptides are disclosed. In some cases, a surface anchor polypeptide is a glycosylphosphatidylinisotol (GPI) anchor domain, which can be synthetic. Kits are also provided for practicing the subject methods.
Type:
Application
Filed:
November 9, 2015
Publication date:
February 8, 2018
Inventors:
Aaron Michael RING, Andrew KRUSE, Aashish MANGLIK
Abstract: Provided are methods and compositions to improve fungal disease resistance and/or nematode resistance in various crop plants. Also provided are combinations of compositions and methods to improve fungal disease resistance and/or nematode resistance in various crop plants.
Type:
Application
Filed:
August 21, 2017
Publication date:
February 8, 2018
Inventors:
Michael J. Crawford, Michelle L. Gasper, Xiangqian Li, Barry J. Shortt, Deryck Jeremy Williams
Abstract: The present invention relates to a method for manipulating the growth and/or structure of a plant, for increasing biomass. The method of the invention is achieved by increasing the expression and/or activity of PXY and/or CLE in the vascular tissue of a plant.
Type:
Application
Filed:
March 1, 2016
Publication date:
February 8, 2018
Inventors:
Simon Turner, Laxmi Mishra, Manoj Kumar, Peter Etchells
Abstract: The present invention provides a wound inducible expression construct and a method of its preparation. The invention provides a methods for isolation of an early wound inducible promoter that is activated within 5 minutes of any form of wounding (mechanical or biological) and a process of making transgenic plants in which expression of GUS/Insecticidal protein is regulated by Promoter (I.D.1) in a wound inducible manner for local expression of a chimeric gene used in this method and plants obtained thereby, and to the process for obtaining resistance to insect feeding.
Type:
Application
Filed:
April 29, 2015
Publication date:
February 8, 2018
Inventors:
Aniruddha Prafullachandra Sane, Saurabh Prakash Pandey, Amar Pal Singh
Abstract: The present invention provides a method for reducing at least one tobacco-specific nitrosamine or a precursor thereof in tobacco comprising modifying the tobacco plant by increasing the activity or expression of a LBD (lateral organ bound domain) nitrogen-responsive transcription factor. The present invention also provides tobacco cells, tobacco plants, tobacco plant propagation materials, harvested leaves, processed tobaccos, or tobacco products obtainable in accordance with the invention.
Type:
Application
Filed:
February 4, 2016
Publication date:
February 8, 2018
Inventors:
PASCALINE LE LAY, JUAN PABLO SANCHEZ TAMBURRINO
Abstract: The invention provides methods for producing a plant with increased stress-tolerance and yield, as well as chimeric genes for use according to the methods and plant comprising such chimeric genes.
Type:
Application
Filed:
October 20, 2017
Publication date:
February 8, 2018
Inventors:
Anna AMTMANN, Matthew HANNAH, Veronique GOSSELE, Manuel LOPEZ-VERNAZA, Giorgio PERRELLA, Christoph VERDUYN
Abstract: The present invention relates to Rubisco activase (RCA) and functionally equivalent polypeptides that confer thermostability to a complex comprising Rubisco activase and Ribulose-1,5-bis-phosphate carboxylase/oxygenase (Rubisco).
Abstract: The present invention relates to isolated nucleic acid molecules and their corresponding encoded polypeptides able confer the trait of improved plant size, vegetative growth, growth rate, seedling vigor and/or biomass in plants challenged with saline and/or oxidative stress conditions. The present invention further relates to the use of these nucleic acid molecules and polypeptides in making transgenic plants, plant cells, plant materials or seeds of a plant having plant size, vegetative growth, growth rate, seedling vigor and/or biomass that are improved in saline and/or oxidative stress conditions with respect to wild-type plants grown under similar conditions.
Type:
Application
Filed:
August 16, 2017
Publication date:
February 8, 2018
Inventors:
Fasong ZHOU, Kenneth A. Feldmann, Julissa Sosa
Abstract: The invention relates to a new gene that produces a protein which is capable of inferring oomycete resistance, more preferably resistance to Phytophthora infestans when expressed in a plant, wherein said nucleotide sequence encodes a protein that is encoded by the protein produced by the nucleotide sequence of FIG. 10 or the protein depicted in FIG. 11 or a nucleotide sequence that codes for a protein that has an identity of at least 95% with said protein produced by the nucleotide sequence of FIG. 10 or the protein depicted in FIG. 11. The invention also relates to a method for providing at least partial resistance or increasing resistance in a plant against an oomycete infection comprising providing a plant or a part thereof with a nucleotide sequence as indicated above or a functional fragment thereof, preferably wherein said plant is a plant from the Solanaceae family, more preferably Solanum tuberosum.
Type:
Application
Filed:
September 17, 2015
Publication date:
February 8, 2018
Applicant:
Wageningen Universiteit
Inventors:
Jacobus Hubertus VOSSEN, Kwang Ryong JO, Richard Gerardus Franciscus VISSER
Abstract: The present invention relates to the field of RNA-mediated gene silencing in insect species. The present invention is based, in part, on the inventors' sequencing of genes from eucalyptus invasive species Gb pest, Glycaspis brimblecombei. In certain aspects, the invention provides Gb nucleic acids, derivatives thereof and the use of such nucleic acids and derivatives as Gb control agents.
Abstract: A method of creating a sperm cell comprising combined genetic material of human males is disclosed. Deoxyribonucleic Acid (DNA) is extracted from a living cell, of a first male. A gene sequence is separated from the Deoxyribonucleic Acid (DNA) of the first male. A recombinant Deoxyribonucleic Acid (DNA) is generated by inserting and attaching the gene sequence of the first male into a Deoxyribonucleic Acid (DNA) of a living cell of a second male. The recombinant Deoxyribonucleic Acid (DNA) is programmed to create Induced pluripotent stem cells (iPSCs). The Induced pluripotent stem cells (iPSCs) are converted into at least one recombinant sperm cell comprising combined genetic material of the first male and the second male.
Abstract: The present invention relates to modified cancer cell lines, wherein the ST3Gal1 glycosyltransferase expression is reduced or inhibited. The use of the modified cancer cell lines with suppressed ST3Gal1 glycosyltransferase to screen a potential cancer therapeutic is provided. The suppression of ST3Gal1 glycosyltransferase expression, with or without an anti-cancer agent is also provided to inhibit the growth of cancer cells in a subject.
Type:
Application
Filed:
August 2, 2017
Publication date:
February 8, 2018
Inventors:
Alice L. Yu, Wen-Der Lin, Jung-Tung Hung, Tan-Chi Fan
Abstract: Compositions and methods are provided for making one or more targeted genetic modifications at a target genomic locus by employing homologous recombination facilitated by single or double-strand break at or near the target genomic locus. Compositions and methods for promoting efficiency of homologous recombination between an LTVEC and a target genomic locus in prokaryotic or eukaryotic cells using engineered nucleases are also provided.
Type:
Application
Filed:
October 24, 2017
Publication date:
February 8, 2018
Applicant:
Regeneron Pharmaceuticals, Inc.
Inventors:
David Frendewey, Wojtek Auerbach, David M. Valenzuela, George D. Yancopoulos, Ka-Man Venus Lai
Abstract: Systems and methods for providing alternative fuel, in particular hydrogen photocatalytically generated by a system comprising photoactive nanoparticles and a nitrogenase cofactor are provided. In one aspect, the system includes a water soluble cadmium selenide nanoparticle (CdSe) surface capped with mercaptosuccinate (CdSe-MSA); and a NafY?FeMo-co complex comprising a NafY protein and an iron-molybdenum cofactor (FeMo-co); wherein the CdSe-MSA and the NafY?FeMo-co complex are present in about 1:1 molar ratio in a CdSe-MSA?NafY?FeMo-co system. In various embodiments, when illuminated, the CdSe-MSA?NafY?FeMo-co system is capable of photocatalytically producing hydrogen gas for an extended period of, e.g., at least 5 hours, at least 10 hours, or at least 90 hours. Methods for making and using the same are also provided.
Abstract: The present invention discloses that by combining different di TPS enzymes of class I and class II different diterpenes may be produced including diterpenes not identified in nature. Surprisingly it is revealed that a di TPS enzyme of class I of one species may be combined with a di TPS enzyme of class II from a different species, resulting in a high diversity of diterpenes, which can be produced.
Type:
Application
Filed:
January 30, 2015
Publication date:
February 8, 2018
Inventors:
Bjorn Hamberger, Birger Lindberg Moller, Johan Andersen-Ranberg, Carl Jorg Bohlmann, Philipp Zerbe, Morten Thrane Nielsen
Abstract: The present invention relates to the production of aromatic hexanols and compositions containing them. In particular provided herein are methods of producing hexenols comprising: a) contacting a hydroperoxide of a polyunsaturated fatty acid with a modified hydroperoxide lyase to form a hexenal; and b) reducing the hexenal to a hexenol in the presence of a hydride donor, a ketoreductase, and a co-factor wherein the contacting and reducing steps are carried out at essentially the same time in the substantial absence of baker's yeast.
Abstract: A combination of an electrochemical device for delivering reducing equivalents to a cell, and engineered metabolic pathways within the cell capable of utilizing the electrochemically provided reducing equivalents is disclosed. Such a combination allows the production of commodity chemicals by fermentation to proceed with increased carbon efficiency.
Type:
Application
Filed:
February 23, 2016
Publication date:
February 8, 2018
Applicant:
Biocheminsights, Inc.
Inventors:
David R. Dodds, William B. Armiger, Mattheos Koffas
Abstract: Provided herein is a process for producing a fermentation product from a lignocellulosic feedstock. The process comprises soaking a lignocellulosic feedstock in an aqueous solution to produce a soaked feedstock. The soaked feedstock is at least partially dewatered and the at least partially dewatered feedstock is subjected to pretreating. The pretreatment chemical is sulfur dioxide, sulfurous acid or a combination thereof and can be added to the process at any stage prior to and/or during pretreatment. The pretreated feedstock composition is fed to an enzymatic hydrolysis conducted at a temperature that is higher than 58° C. The cellulose in the pretreated feedstock composition is hydrolyzed with cellulase enzymes in the presence of the dissolved solids to produce glucose. The glucose is fermented to produce the fermentation product.
Abstract: The present invention relates to methods for the enzymatic deacetylation of mannosylerythritol lipids produced by fermentation using lipases. More in particular, the present invention relates to a method for the enzymatic deacetylation of mannosylerythritol lipids using a hydrolyzing enzyme in an organic solvent containing only low amounts of water, preferably, no water. It further provides the use of organic solvents, containing only low amounts of water, more preferably, no water, for the enzymatic deacetylation of mannosylerythritol lipids.
Type:
Application
Filed:
March 1, 2016
Publication date:
February 8, 2018
Applicant:
Universiteit Antwerpen
Inventors:
Eliane Yvonne Goossens, Marc Victor Henri Wijnants
Abstract: The present disclosure relates to the biosynthesis of indigoid dye precursors and their conversion to indigoid dyes. Specifically, the present disclosure relates to methods of using polypeptides to produce indigoid dye precursors from indole feed compounds, and the use of the indigoid dye precursors to produce indigoid dyes.
Type:
Application
Filed:
March 3, 2016
Publication date:
February 8, 2018
Applicant:
The Regents of the University of California
Inventors:
John Eugene DUEBER, Zachary Nicholas RUSS, Tammy Melody HSU, Terry Don JOHNSON, Jr., Bernardo CERVANTES, Ramya Lakshmi PRATHURI, Shyam Pravin BHAKTA, Arthur Muir FONG, III, Luke Nathaniel LATIMER
Abstract: A novel technique for improving secretory production of a heterologous protein by coryneform bacteria is described, as well as a method for secretory production of a heterologous protein. A coryneform bacterium is described that has been modified to have a specific mutation so as to harbor a phoS gene, and when cultured, is able to to produce a heterologous protein by secretory production.
Abstract: Provided are improved fungal strains and use thereof, wherein the fungal strains are capable of producing an altered level of proteins, enzymes, variants and other substances of interest.
Type:
Application
Filed:
February 9, 2016
Publication date:
February 8, 2018
Applicant:
DANISCO US INC.
Inventors:
Elizabeth A. BODIE, Roman RABINOVICH, Aleksandra VIRAG, Michael WARD, Rochelle NGUYEN, James A. SWEIGARD
Abstract: An antibody has at a heavy chain thereof a C-terminal extension that includes at least one glutamine that is a substrate for transglutaminase, enabling the transglutaminase-mediated preparation antibody-drug conjugates using such antibody.
Abstract: The invention is the products and methods associated with purifying overexpressed recombinant recombinases from a host cell line resulting in an un-tagged protein of interest without any additional, non-native amino acids. The invention employs at least one DNA vector that co-expresses a tagged fusion protein and the recomibinase protein with the recombinase protein having an affinity for binding to the the tagged fusion protein. Isolation methods of the recominbase protein include the targeting of the tagged fusion protein.
Abstract: A method for detecting lactic acid and/or acetic acid bacteria in a food-processing matrix comprising a microbial flora (the microbial flora comprising a lactic acid and/or acetic acid bacterial flora) and a fungal flora, the bacterial flora containing an adenosine triphosphate of bacterial origin, the fungal flora containing an adenosine triphosphate of fungal origin, which comprises the following steps: applying, to the matrix, an antifungal having an antifungal action which is lethal, at a first time limit, on the fungal flora, and an antibiotic action which is non lethal, at a second time limit after the first time limit, on the bacterial flora, detecting the microbial flora between the first time limit and the second time limit; in which the lethal antifungal action releases, into the matrix, for the first time limit, adenosine triphosphate of fungal origin and in which the microbial flora is detected between the first time limit and the second time limit by means of the following steps: removing the
Abstract: Nanofluidic devices and methods of the invention are capable of autonomously isolating individual microbial cells using constrictive channels and growing monocultures of the cells for automated characterization of their biochemical properties and interactions with mammalian cells. Single microbial cells, such as bacterial cells, are isolated directly from environmental sources and cultured using chemical factors from their native environment.
Abstract: Disclosed are methods to measure the extent of viral infection and methods to measure latent viral reservoir in cells harboring or suspected of harboring such a reservoir.
Type:
Application
Filed:
August 1, 2017
Publication date:
February 8, 2018
Inventors:
Nathan Mark Sherer, Jordan Thomas Becker
Abstract: Devices, methods, and kits for easy, accurate, and fast estimation of soil microbial load are provided. The devices include membranes that are rapidly wettable with aqueous fluids and retain soil microorganisms of up to 200 micrometers in size, without retaining soil pigments. The method requires reconstituting a soil sample in an extraction fluid so that the microorganisms in the soil sample are released from particles of the soil sample and into the extraction fluid. The method does not require a filtration step. The extraction fluid containing the microorganisms suspended therein is then pipetted onto the membrane device, or the membrane is dipped into the extraction fluid. The color appearing on the membrane is then compared to color intensity gray scale strips to determine the soil microbial load.
Abstract: A method of quantifying ammonia, the method includes performing a first reaction in which a test liquid containing ammonia is reacted with adenosine triphosphate and L-glutamic acid in the presence of glutamine synthetase to produce phosphoric acid, performing a second reaction in which the produced phosphoric acid is reacted with pyruvic acid in the presence of pyruvate oxidase, and measuring a component consumed or produced by the second reaction, to quantify ammonia, wherein a reaction to produce adenosine triphosphate from adenosine diphosphate mediated by pyruvate kinase is not carried out.
Type:
Application
Filed:
August 2, 2017
Publication date:
February 8, 2018
Applicant:
ARKRAY, Inc.
Inventors:
Koji Katsuki, Takehiro Funamoto, Masaki Murakami
Abstract: A sampler and a method of parameterization by calibration of digital circuits and non-invasive determination of the concentration of several biomarkers simultaneously and in real time. The method makes use of equipment which, from a set of luminous signatures—spectrum—provided by a spectrophotometer (E5) (E6), applies a digital filter that breaks down the spectrum into sub-spectra that shows the digital signatures of relevant markers and, through a digital decoder, the concentration of a set of several biomarkers is obtained simultaneously and in real time.
Type:
Application
Filed:
August 3, 2017
Publication date:
February 8, 2018
Inventors:
José António Martins, Marcelo Adorni Pereira, Vanderlei Pereira Ferreira
Abstract: The purpose of the invention is to provide a nucleic acid analyzer that sets and executes temperature regulation, said temperature regulation matching the characteristics of analyzing items and the configuration of the analyzer, by a simple operation while preventing deterioration in analytical performance caused by partial overheating of a liquid reaction mixture to thereby improve temperature change speed and shorten analysis time. To achieve the above purpose, provided is a method wherein, in the case of performing overshooting: as a first processing, the temperature is continuously elevated until reaching a target overshoot temperature; as a second processing, after reaching the aforesaid temperature, the overshoot target temperature is maintained for a preset period of time; and, as a third processing, the temperature is continuously lowered until reaching a target temperature of the liquid reaction mixture.
Abstract: A chemically-enhanced primer is provided comprising a negatively charged moiety (NCM), an oligonucleotide sequence having a) non-nuclease resistant inter-nucleotide linkages or b) at least one nuclease resistance inter-nucleotide linkage. The chemically-enhanced primer can be used for sequencing and fragment analysis. Methods for synthesizing the chemically-enhanced primer as well as a method of preparing DNA for sequencing, a method of sequencing DNA, and kits containing the chemically-enhanced primer are also provided. The method of sequencing DNA can comprise contacting amplification reaction products with the composition wherein excess amplification primer is degraded by the nuclease and the chemically-enhanced primer is essentially non-degraded.
Abstract: Embodiments of the present invention are directed toward devices, systems, and methods for purifying nucleic acids to conduct polymerase chain reaction (PCR) assays. In one example, a method includes generating complexes of silica beads and nucleic acids in a lysis buffer, transporting the complexes through an immiscible fluid to remove interfering compounds from the complexes, further transporting the complexes into a density medium containing components required for PCR where the nucleic acids disassociate from the silica beads, and thermocycling the contents of the density medium to achieve PCR. Signal may be detected from labeling agents in the components required for PCR.
Type:
Application
Filed:
September 27, 2017
Publication date:
February 8, 2018
Inventors:
Chung-Yan Koh, Yooli Kim Light, Matthew Ernest Piccini, Anup K. Singh
Abstract: A heating mechanism for use in DNA applications such as DNA amplification, extraction and sterilization is provided. Nanoparticles having photo-thermal properties are put in contact with a reaction mixture and irradiated with an activation light beam to activate these photo-thermal properties, thereby releasing heat. Nanoparticles of several types may be used. Use of the same nanoparticles or of different one to monitor the reaction using a different light beam is also presented.
Type:
Application
Filed:
September 29, 2017
Publication date:
February 8, 2018
Inventors:
Philip ROCHE, Andrew KIRK, Lenore BEITEL, Miltiadis PALIOURAS, Mark TRIFIRO, Vamsy CHODAVARAPU, Mohamed NAJIH, Joachim THIEMANN
Abstract: The subject matter of the present invention is a microfluidic process for treating and analysing a solution containing a biological material, comprising a step of introducing the solution into microchannels of a microfluidic circuit (1), a step of forming drops of this solution, under the effect of modifications of the surface tension of the solution, a step of moving the drops to one or more drop storage zones(s) (130), under the effect of modifications of the surface tension of the drops, a step of treating the drops and a step of analysing the drops.
Type:
Application
Filed:
October 18, 2017
Publication date:
February 8, 2018
Inventors:
Charles Baroud, Remi Dangla, Paul Abbyad, Silvan Turkcan
Abstract: Transposon nucleic acids comprising a transposon end sequence and a calibration sequence for DNA sequencing in the transposon end sequence. In one embodiment, the transposon end sequence is a Mu transposon end. A method for the generation of DNA fragmentation library based on a transposition reaction in the presence of a transposon end with the calibration sequence providing facilitated downstream handling of the produced DNA fragments, e.g., in the generation of sequencing templates.
Type:
Application
Filed:
October 24, 2017
Publication date:
February 8, 2018
Inventors:
Ian Kavanagh, Laura-Leena Kiiskinen, Heli Haakana
Abstract: The invention relates to a method for quantitative monitoring of bacterial endospores in an aqueous environment of a paper or board mill. The method comprises at least the following steps: obtaining at least a first aqueous sample originating from the industrial aqueous environment; destroying bacteria in vegetative form in the first sample by a suitable treatment, preferably by heating the first sample to a desired temperature; adding intercalating agent (such as PMA) to the treated first sample and allowing it to interact (e.g. by cross-linking) with the destroyed bacteria, so that the nucleic acid from the destroyed bacteria are unavailable for PCR; and determining the endospore level in the first sample by using quantitative polymerase chain reaction (qPCR) in which only the DNA from the endospores is available for amplification.
Type:
Application
Filed:
February 26, 2016
Publication date:
February 8, 2018
Applicant:
Kemira Oyj
Inventors:
Kalle Riihinen, Marko Lauraeus, Marko Kolari, Juhana Ahola
Abstract: The invention provides monoferrocenyl compounds of general formula I. The invention also provides substrates labelled with the compounds, functionalised derivatives of the compounds and methods of using the compounds, functionalised derivatives and labelled substrates in electrochemical assays.
Type:
Application
Filed:
October 13, 2017
Publication date:
February 8, 2018
Inventors:
Barrie J. Marsh, Christopher G. Frost, Jonathan Sharp
Abstract: Disclosed is a method for determining the presence of Mycobacterium avium complex nucleic acids in a biological sample. In particular, the mig gene of M. avium and the DT1 gene of M. intracellulare are detected, preferably following amplification. In addition, the method distinguishes between species of M. avium and M. intracellulare.
Abstract: The present disclosure relates, in some aspects, to the field of nucleic acid detection. Disclosed herein are methods and compositions for detecting nucleic acids using synthetic single-stranded ribonucleic acids (RNAs). In certain embodiments, synthetic single-stranded RNAs are used to detect therapeutic nucleic acids, such as therapeutic deoxyribonucleic acids (DNAs) and/or therapeutic ribonucleic acids (RNAs).
Abstract: The present invention provides compositions and methods for assaying the activity of nicking enzyme and polymerase in a reaction involving the use of a nucleic acid substrate molecule that detects nicking enzyme and polymerase extension activities by the release of a detectable reporter (e.g., a fluorophore).
Abstract: A method for detecting a genetic mutation is provided. The method comprising: carrying out PCR using a template DNA, a forward primer, a reverse primer and a wild-type oligonucleotide; and detecting an amplified DNA. In the method, the wild-type oligonucleotide comprises LNA or BNA, and on a DNA strand of the template, with which the wild-type oligonucleotide is hybridized, a region with which the wild-type oligonucleotide is a hybridized and a region with which the forward primer or the reverse primer is hybridized partially overlap each other, or the regions are separated from each other by 1 to 18 bases.
Abstract: The present disclosure relates to compositions, methods and kits for quantitative analysis of a plurality of nucleic acid target molecules in a sample. In some embodiments, the methods comprise associating molecular label sequences with target nucleic acid molecules without an enzymatic reaction.
Abstract: The present invention relates to an in vitro cell-free process for production of deoxyribonucleotides (DNAs) comprising at least one hairpin, corresponding DNA products and uses thereof, and oligonucleotides and kits useful in the process of the invention.
Type:
Application
Filed:
February 17, 2016
Publication date:
February 8, 2018
Inventors:
Paul ROTHWELL, Neil PORTER, Lydia VINEY, Thomas ADIE
Abstract: Present invention disclosed a novel method for detecting and quantifying target DNA from the biological sample. It provides a method of amplification of DNA sequences with loop-mediated isothermal amplification (LAMP) and its easy and accurate detection and quantification by electrochemiluminescence (ECL) technique. The target LAMP DNA is bound electrostatically with [Ru(bpy)3]+2 on the carbon electrode surface, and an ECL reaction is triggered by tripropylamine (TPrA) to yield luminescence. This is a highly sensitive strategy for the detection of sequence-specific DNA from different biological samples at picogram levels. The target DNA of Sus scrofa (pork) meat was detected as low as 1 pg/?L (3.43×10?1 copies/?L) and for Bacillus subtilis DNA samples the detection limit of 10 pg/?L (2.2×103 copies/?L) was achieved.
Abstract: Improved methods for the detection and quantitation of a target nucleic acid in a sample using a non-extending helper oligonucleotide are described. The methods include contacting nucleic acids in a sample with amplification reagents including one or more primers, one or more non-extending helper oligonucleotides, and one or more probes. The non-extending helper oligonucleotide facilitates and increases the target nucleic acid accessibility of one or more of the primers, result in greater accumulation of amplicon production, thereby increasing the efficiency and sensitivity of the amplification assay, including amplification assays for Hepatitis C Virus (HCV), for example, HCV Genotype 5. Kits, articles of manufacture, and reaction mixtures are also provided.