Showing papers on "Nucleic acid methods published in 2016"

Recent Advances in Developing Small Molecules Targeting Nucleic Acid

Abstract: Nucleic acids participate in a large number of biological processes. However, current approaches for small molecules targeting protein are incompatible with nucleic acids. On the other hand, the lack of crystallization of nucleic acid is the limiting factor for nucleic acid drug design. Because of the improvements in crystallization in recent years, a great many structures of nucleic acids have been reported, providing basic information for nucleic acid drug discovery. This review focuses on the discovery and development of small molecules targeting nucleic acids.

Nucleic acid sequence analysis from single cells

Abstract: Presented herein are methods and compositions for multiplexed single cell gene expression analysis. Some methods and compositions include the use of droplets and/or beads bearing unique barcodes such as unique molecular barcodes (UMI).

Nucleic acid tool enzymes-aided signal amplification strategy for biochemical analysis: status and challenges

Abstract: Owing to their highly efficient catalytic effects and substrate specificity, the nucleic acid tool enzymes are applied as 'nano-tools' for manipulating different nucleic acid substrates both in the test-tube and in living organisms. In addition to the function as molecular scissors and molecular glue in genetic engineering, the application of nucleic acid tool enzymes in biochemical analysis has also been extensively developed in the past few decades. Used as amplifying labels for biorecognition events, the nucleic acid tool enzymes are mainly applied in nucleic acids amplification sensing, as well as the amplification sensing of biorelated variations of nucleic acids. With the introduction of aptamers, which can bind different target molecules, the nucleic acid tool enzymes-aided signal amplification strategies can also be used to sense non-nucleic targets (e.g., ions, small molecules, proteins, and cells). This review describes and discusses the amplification strategies of nucleic acid tool enzymes-aided biosensors for biochemical analysis applications. Various analytes, including nucleic acids, ions, small molecules, proteins, and cells, are reviewed briefly. This work also addresses the future trends and outlooks for signal amplification in nucleic acid tool enzymes-aided biosensors.

Methods for studying nucleic acids

Abstract: The present invention provides a novel method for preparing a sequencing library and studying molecular interactions involving a nucleic acid. In particular, the invention relates to a method for preparing a sequencing library, the method comprising the addition of an agent binding to chromatin to a sample comprising a nucleic acid; isolating chromatin bound by said agent; addition of transposase to the isolated chromatin; isolating nucleic acid from chromatin; and obtaining a sequencing library. Moreover, the present invention relates to a method for mapping of molecular interactions involving a nucleic acid, the method comprising the addition of an agent binding to chromatin to a sample comprising a nucleic acid; isolating chromatin bound by said agent; addition of transposase to the isolated chromatin; isolating nucleic acid from chromatin; amplification of nucleic acid; sequencing of amplified nucleic acid; and identifying molecular interactions.

Engineered nucleic acid-targeting nucleic acids

Abstract: The present disclosure provides engineered polynucleotide sequences that form scaffolds and nucleoprotein complexes comprising such engineered polynucleotide sequences that form scaffolds and nucleic acid binding proteins Nucleic acid sequences encoding the engineered polynucleotide sequences that form scaffolds, as well as expression cassettes, vectors and cells comprising such polynucleotide sequences, are described A variety of methods for making and using the engineered polynucleotide sequences that form scaffolds are also disclosed

Capture of nucleic acids using a nucleic acid-guided nuclease-based system

Abstract: Provided herein are methods and compositions for the capture of nucleic acids, for example by using a nucleic acid-guided nuclease-based system.

Methods and compositions for targeted nucleic acid sequence coverage

Abstract: The present invention is directed to methods, compositions and systems for analyzing sequence information from targeted regions of a genome. Such targeted regions may include regions of the genome that are poorly characterized, highly polymorphic, or divergent from reference genome sequences.

Nucleic Acids In Chemistry And Biology

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Combinatorial sets of nucleic acid barcodes for analysis of nucleic acids associated with single cells

Abstract: Provided herein are methods of identifying the origin of a nucleic acid sample. The methods include forming a reaction mixture comprising a nucleic acid sample comprising nucleic acid molecules from a single cell and a set of barcodes, incorporating the set of barcodes into the nucleic acid molecules of the sample, and identifying the set of barcodes incorporated into the nucleic acid molecules of the single cell thereby identifying the origin of the nucleic acid sample.

Submicrometer‐Sized Thermometer Particles Exploiting Selective Nucleic Acid Stability

Abstract: Encapsulated nucleic acid selective damage quantification by real-time polymerase chain reaction is used as sensing mechanism to build a novel class of submicrometer size thermometer. Thanks to the high thermal and chemical stability, and the capability of storing the read accumulated thermal history, the sensor overcomes some of current limitations in small scale thermometry.

Crispr-cas compositions and methods

Abstract: Class 2 CRISPR-Cas nucleoprotein complexes are disclosed comprising a Class 2 CRISPR-Cas protein, a CRISPR-Cas associated polynucleotide lacking a spacer element (casPN), and a distinct spacer element sequence polynucleotide (sesPN) comprising a target nucleic acid binding sequence. These complexes are capable of site-directed binding to a target nucleic acid complementary to the target nucleic acid binding sequence of the sesPN. The Class 2 CRISPR-Cas nucleoprotein complexes facilitate site-specific modifications, including cleavage and mutagenesis, of a target nucleic acid sequence. Polynucleotide sequences, expression cassettes, vectors, compositions, and kits for carrying out a variety of methods are also described. Furthermore, the present specification provides guidance for methods of regulating expression of a target nucleic acid sequence, production of genetically modified cells, compositions of modified cells, transgenic organisms, pharmaceutical compositions, as well as a variety of other compositions and methods involving the Class 2 CRISPR-Cas nucleoprotein complexes comprising casPNs, sesPNs, and Cas proteins.

Compositions and methods for target nucleic acid modification

Abstract: The present disclosure provides a complex comprising a nanoparticle; a Type II or a Type V CRISPR system comprising a site -directed DNA-modifying polypeptide and a guide RNA; and a polycation-based endosomal escape polymer. The present disclosure provides methods of making and using a complex of the present disclosure.

Application of nucleic acid analogues as receptor layers for biosensors

Abstract: Nucleic acid-based biosensors are typically used to detect DNA or RNA fragments of clinical importance. Moreover, these sensors can also be used to detect different analytes, including viruses, peptides, small organic molecules and metal ions. However, the lack of resistance to restriction enzymes prevents the use of these sensors in in vivo measurements. Moreover, due to their reduced stability under environmental conditions as well as non-specific interactions, the usefulness of DNA biosensors can be limited. To mitigate these problems, analogues of nucleic acids can be used. These compounds are structurally similar to natural DNA, with alternative backbones. Due to their excellent affinity and specificity toward complementary strands, the most useful and popular nucleic acid analogues are peptide nucleic acid (PNA), locked nucleic acid (LNA) and phosphorothioate oligonucleotide (PTO). Herein, we present an overview of biosensors employing nucleic acid analogues as receptor layers, published in the last few years. Their advantages and their limitations are discussed.

Modified Nucleic Acids in Biology and Medicine

Abstract: Transfer RNAs (tRNAs) are essential components of the protein translation machinery. In order to become fully active, they need to be heavily modified post-transcriptionally. Such modifications affect the structure, stability and functionality of tRNAs; however, their exact roles at the molecular level remain largely elusive. Here we focus on the biological functions of tRNA modifications associated to human diseases and how such information can be used for biomedical applications. We put an emphasis on mitochondrial-linked dysfunctions, metabolic disorders, neurological defects and cancer. We also present methods and approaches currently used in the clinic to detect and monitor different human A.G. Torres Institute for Research in Biomedicine (IRB Barcelona), Barcelona Institute for Science and Technology, C/Baldiri Reixac 10, Barcelona 08028, Catalonia, Spain e-mail: adriangabriel.torres@irbbarcelona.org L. Ribas de Pouplana (*) Institute for Research in Biomedicine (IRB Barcelona), Barcelona Institute for Science and Technology, C/Baldiri Reixac 10, Barcelona 08028, Catalonia, Spain Catalan Institution for Research and Advanced Studies (ICREA), P/Lluis Companys 23, Barcelona 08010, Catalonia, Spain e-mail: lluis.ribas@irbbarcelona.org © Springer International Publishing Switzerland 2016 S. Jurga et al. (eds.), Modified Nucleic Acids in Biology and Medicine, RNA Technologies, DOI 10.1007/978-3-319-34175-0_1 1 pathologies involving tRNA modifications or tRNA modification enzymes, and, additionally, we propose novel tRNA modification-based strategies that could be used for diagnosis, prognosis or treatment of human diseases.

Nucleic Acids And Molecular Biology

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Multiple-emulsion nucleic acid amplification

Abstract: Multiple-emulsion nucleic acid amplification allows nucleic acids contained in biological systems to be detected, quantitated and/or sorted based on their sequence as detected with nucleic acid amplification techniques, e.g., PCR. The nucleic acids can be free floating or contained within living or nonliving structures, including particles, viruses, and cells. The nucleic acids can include, e.g., DNA or RNA. Systems and devices for use in practicing methods of the disclosure are also provided.

Methods for sequencing nucleic acids

Abstract: Provided herein are methods and compositions for the sequencing of long nucleic acids, such as DNA. The methods and compositions are suited for the spatial labeling and sequencing of long nucleic acid molecules.

Nucleic Acid Aptamers

Abstract: Optical nanosensors are based on particles with diameters from 20 to 200 nm containing sensory elements. The latter are comprised of one or more signaling molecules and one or more references, which allow measurements to be ratiometric and hence independent on the amount of sensor. The signaling molecules may range from simple ion-binding fl uorophores, e.g., pH-sensitive dyes, to complex biochemical assays. Aptamers are ideal for use in nanosensors because they are relatively easy to modify chemically and hence to transform into signaling molecules, and their binding affi nities may be fi ne-tuned to a desired measuring range in the selection process. Here we fi rst describe the selection of metabolite binding aptamers, how they are transformed into signaling molecules using a molecular beacon construct and then how they are inserted into nanoparticles. Finally, we briefl y describe how the sensors are calibrated before inserted into cells to measure metabolite concentration in real time. As examples we present aptamers binding to key metabolites in cells: ATP and fructose 1, 6-bisphosphate (FBP).

Optimized nucleic acid molecules

Abstract: The present invention provides optimized nucleic acid molecules, methods for optimization of nucleic acid molecules and uses of optimized nucleic acid molecules. A modular design principle is provided that is suitable to generate a nucleic acid, particularly mRNA, which is tailored for a respective application. The nucleic acid molecules of the present invention can be obtained by the versatile combination of multiple modules on nucleic acid level. Such nucleic acid, e.g. mRNA, can be tailored by combining one or more modules, comprising (i) a nucleic acid moiety encoding a polypeptide of interest (e.g. a protein potentially producing a therapeutic outcome) and (ii) at least one further coding or non-coding nucleic acid moiety, e.g. selected among nucleic acid moieties encoding a polypeptide element, such as a secretory signal peptide (SSP), a multimerization element (dimerization, trimerization, tetramerization and oligomerization), a virus like particle (VLP) forming element, a transmembrane element, a dendritic cell targeting element, an immunological adjuvant element, an element promoting antigen presentation; a 2A peptide; a peptide linker element, elements that extend protein half-life, and/or any other polypeptide or protein. Non-coding nucleic acid moieties may be selected e.g. from the group comprising 3'-UTR, 5'-UTR, IRES element, miRNA moiety, histone stem loop, poly(C) sequence, polyadenylation signal, polyA-sequence. The optimized nucleic acid molecule can further be characterized by the presence of at least one modified nucleoside. The versatility of the present invention allows for rational design of a large variety of different nucleic acid molecules with desired properties.

A functionally integrated thermoplastic microdevice for one-step solid-phase-based nucleic acid purification and isothermal amplification for facile detection of foodborne pathogen.

Abstract: In this study, we fabricate a functionally integrated monolithic thermoplastic microdevice for continuous operation of nucleic acid purification and amplification using polycarbonate (PC). A solid-phase-based purification and subsequent isothermal amplification, specifically, thermal helicase-dependent amplification (tHDA), was performed in a single operation in a valve-free manner. PC microdevice was assembled using modified thermal bonding process under relatively low temperature and pressure condition, realized by surface chemical modification of PC into hydrophilic property using amine-bearing polyethyleneimine (PEI). After the device sealing, only the microchannel parts were selectively modified to be hydrophobic, using epoxy-terminated poly(dimethylsiloxane) (PDMS) (epoxy-PDMS) on amine-coated surface for stepwise introduction of multiple reagents in a valve-free manner. Using the integrated PC microdevice, nucleic acids from genetically modified Escherichia coli (E. coli) O157:H7 were captured inside a chamber bearing amine functionality, by electrostatic interaction, and were subsequently amplified isothermally in the same chamber. Purified DNA captured inside the microchamber was detected directly inside the chamber by fluorescence measurement, and a 92-bp long EaeA gene, inserted into the E. coli O157:H7, was successfully amplified using the integrated PC microdevice in less than 90 min, paving the way for facile identification of foodborne pathogens with simple operation and reduced peripheral operations applicable for portable healthcare purposes. Biotechnol. Bioeng. 2016;113: 2614-2623. © 2016 Wiley Periodicals, Inc.

DNA sulfite conversion and purification method

Abstract: The invention discloses a DNA sulfite conversion and purification method. The method does need early sodium hydroxide denaturating treatment conducted on nucleic acid, adopts varying-temperature conversion conditions different from a commercialized reagent on the market and can perform a conversion experiment under the constant temperature condition. A magnetic bead method is adopted to perform nucleic acid purification after nucleic acid methylation conversion, a high-salt low pH value is utilized to perform nucleic acid separation and purification, then a low-salt high pH value is utilized to perform elution, the step of performing desulfonating by adopting sodium hydroxide on the market at present is omitted in the purification process, and meanwhile high-conversion-rate, high-quality and high-purity DNA can be obtained only by adopting a one-step washing step. The DNA sulfite conversion and purification method has the advantages of being quick, simple and convenient to operate and improving the conversion efficiency, the extraction efficiency and the extraction purity.

A Rapid and Automated Device for Purifying Nucleic Acids.

Abstract: We have developed a rapid, automated nucleic acid purification device in a single cartridge containing silica-coated magnetic beads. We succeeded in extracting the matrix protein gene of influenza A virus from pharyngeal swab samples within 3 min. The device will be widely applicable to detect a specific gene from the various samples for clinical diagnosis and genetic research.

Field-effect apparatus and methods for sequencing nucelic acids

Abstract: The present disclosure provides a method for sequencing nucleic acids. The method can include polymerase catalyzed incorporation of nucleotides into a nascent nucleic acid strand against a nucleic acid template, wherein the polymerase is attached to a charge sensor that detects nucleotide incorporation events. One or more non-natural nucleotide types that each produce a unique signatures at the charge sensor can be used to uniquely identify different nucleotides in the template nucleic acid.

Nucleic acid sample enrichment

Abstract: This invention relates to the preparation of nucleic acid samples for analysis and certain methods and tools for the selection of specific regions of interest from a nucleic acid sample. The method is particular advantageous for multiplexed amplification reactions, whereby one fixed primer can be used. The method can be used in the preparation of enriched libraries from single stranded, highly fragmented samples.

A powerless on-the-spot detection protocol for transgenic crops within 30 min, from leaf sampling up to results

Abstract: The requirement of power-dependent instruments or excessive operation time usually restricts current nucleic acid amplification methods from being used for detection of transgenic crops in the field. In this paper, an easy and rapid detection method which requires no electricity supply has been developed. The time-consuming process of nucleic acid purification is omitted in this method. DNA solution obtained from leaves with 0.5 M sodium hydroxide (NaOH) can be used for loop-mediated isothermal amplification (LAMP) only after simple dilution. Traditional instruments like a polymerase chain reaction (PCR) amplifier and water bath used for DNA amplification are abandoned. Three kinds of dewar flasks were tested and it turned out that the common dewar flask was the best. Combined with visual detection of LAMP amplicons by phosphate (Pi)-induced coloration reaction, the whole process of detection of transgenic crops via genetically pure material (leaf material of one plant) could be accomplished within 30 min. The feasibility of this method was also verified by analysis of practical samples.

Rapid Methods for the Extraction of Nucleic Acids from Biological Samples

Abstract: The invention is directed to compositions and methods for rapidly and efficiently extracting nucleic acids and/or targeted nucleic acids sequences from biological samples. The methods of the invention comprise combining the sample with a buffer and magnetic silicon beads and concentrating the beads with a magnet or other electrical field. Liquid may be removed, or not, and an alkaline buffer is added followed by magnetic carboxy beads in a binding buffer so that nucleic acids transfer to the carboxy beads, which can be easily and quickly isolated once again with a magnet. Total nucleic acid extraction is greatly enhanced. Extracted nucleic acids can be analyzed, for example, by PCR wherein the nucleic acids can be identified and characterized. Carboxy beads may also contain a ligand so as to target specific nucleic acid sequences. The invention is also directed to kits comprising the tools and compositions for performing the methods of the invention.