1679 results about "Double stranded" patented technology

The present invention provides methods, compositions and kits for using a transposase and a transposon end for generating extensive fragmentation and 5′-tagging of double-stranded target DNA in vitro, then using a DNA polymerase for generating 5′- and 3′-tagged single-stranded DNA fragments without performing a PCR amplification reaction, wherein the first tag on the 5′-ends exhibits the sequence of the transferred transposon end and optionally, an additional arbitrary sequence, and the second tag on the 3′-ends exhibits a different sequence from the sequence exhibited by the first tag. The method is useful for generating 5′- and 3′-tagged DNA fragments for use in a variety of processes, including processes for metagenomic analysis of DNA in environmental samples, copy number variation (CNV) analysis of DNA, and comparative genomic sequencing (CGS), including massively parallel DNA sequencing (so-called “next-generation sequencing.)

In a first aspect, the present invention provides methods for authenticating a nucleic acid molecule and its sequence with a molecular barcode and batch-stamp. In another aspect, the present invention provides methods for authenticating a nucleic acid amplification product. In a further aspect, the present invention provides compositions for encoding both single-stranded and double-stranded target nucleic acids with coded oligonucleotides. The compositions are useful in the practice of the methods of the invention.

This disclosure describe three related novel methods for Recombinase-Polymerase Amplification (RPA) of a target DNA that exploit the properties of the bacterial RecA and related proteins, to invade double-stranded DNA with single stranded homologous DNA permitting sequence specific priming of DNA polymerase reactions. The disclosed methods has the advantage of not requiring thermocycling or thermophilic enzymes. Further, the improved processivity of the disclosed methods allow amplification of DNA up to hundreds of megabases in length.

A method is provided for adding a terminal sequence tag to nucleic acid molecules for use in RNA or DNA amplification. The method involves contacting with a mixture of oligonucleotides, each having a sequence tag template, a random sequence and a blocked 3′ terminus, under conditions such that, the random sequence anneals with the nucleic acid molecules and the nucleic acid molecules are extended using the sequence tag template as template. For synthesis of RNA from DNA molecules having terminal sequence tags, the method includes forming DNA templates having a double stranded promoter sequence and synthesizing RNA from the DNA templates. For amplification of sequences from DNA molecules having terminal sequence tags, the method includes forming DNA templates by extension of one primer having a sequence that is complementary to the terminal sequence tag and another primer having a sequence that is derived form one of the DNA molecules.

One aspect of the present invention relates to a double-stranded oligonucleotide comprising at least one aralkyl ligand. In certain embodiments, an aralkyl ligand is bound to only one of the two oligonucleotide strands comprising the double-stranded oligonucleotide. In certain embodiments, an aralkyl ligand is bound to both of the oligonucleotide strands comprising the double-stranded oligonucleotide. In certain embodiments, the oligonucleotide strands comprise at least one modified sugar moiety. In certain embodiments, at least one phosphate linkage in the oligonucleotide has been replaced with a phosphorothioate linkage. In a preferred embodiment, the aralkyl ligand is naproxen or ibuprofen. Another aspect of the present invention relates to a single-stranded oligonucleotide comprising at least one aralkyl ligand. In certain embodiments, the oligonucleotide comprises at least one modified sugar moiety. In certain embodiments, at least one phosphate linkage in the oligonucleotide has been replaced with a phosphorothioate linkage. In a preferred embodiment, the aralkyl ligand is naproxen or ibuprofen. The aralkyl ligand improves the pharmacokinetic properties of the oligonucleotide.

A method of isolating genomic or RNA-derived duplex fragments which are unique to one of two fragment mixtures. The fragments in positive-source and negative-source mixtures are separately equipped with end linkers, and each mixture is amplified by successive primed-strand replications, using a single primer which is homologous to the associated linker. The second-source linker is biotinylated, and the fragments in this mixture are hybridized in molar excess with the fragments in the positive-source mixture. DNA species which are not hybridized with the biotinylated species, i.e., species that are unique to the positive-source mixture, are isolated after removal of hybridized species by affinity chromatography. Also disclosed is a method of amplifying a mixture of DNA fragments by repeated linker/primer replication.

A method for isolating nucleic acid which comprises:

• • (a) applying a sample comprising cells containing nucleic acid to a filter, whereby the cells are retained as a retentate and contaminants are removed;
  • (b) lysing the retentate from step (a) while the retentate is retained by the filter to form a cell lysate containing the nucleic acid;
  • (c) filtering the cell lysate with the filter to retain the nucleic acid and remove remaining cell lysate;
  • (d) optionally washing the nucleic acid retained by the filter; and
  • (e) eluting the nucleic acid, wherein the filter composition and dimensions are selected so that the filter is capable of retaining the cells and the nucleic acid.

Additionally, there is provided a substrate for lysing cells and purifying nucleic acid having a matrix and a coating and an integrity maintainer for maintaining the purified nucleic acid. Also provided is a method of purifying nucleic acid by applying a nucleic acid sample to a substrate having an anionic detergent affixed to a matrix, the substrate physically capturing the nucleic acid, bonding the nucleic acid to a substrate and generating a signal when the nucleic acid bonds to the substrate indicating the presence of the nucleic acid. A kit for purifying nucleic acid containing a coated matrix and an integrity maintenance provider for preserving the matrix and purifying nucleic acid is also provided. Further, there is provided a method for quantifying DNA, such as double-stranded or genomic DNA, isolated from cells, such as leukocytes to determine the numbers of leukocytes in a sample of leukoreduced blood.

The invention relates to a double-stranded compound, preferably an oligoribonucleotide, which down-regulates the expression of a human p53 gene. The invention also relates to a pharmaceutical composition comprising the compound, or a vector capable of expressing the oligoribonucleotide compound, and a pharmaceutically acceptable carrier. The present invention also contemplates a method of treating a patient suffering from alopecia or acute renal failure or other diseases comprising administering to the patient the pharmaceutical composition in a therapeutically effective dose so as to thereby treat the patient. The alopecia may be induced by chemotherapy or radiotherapy, and the patient may be suffering from cancer, in particular breast cancer.

The present invention relates to a double-stranded ribonucleic acid (dsRNA) having a nucleotide sequence which is substantially identical to at least a part of a target gene and which is no more than 49, preferably less than 25, nucleotides in length, and which comprises a complementary (antisense) RNA strand having a 1 to 4 nucleotide overhang at the 3′-end and a blunt 5′-end. The invention further relates to a pharmaceutical composition comprising the dsRNA and a pharmaceutically acceptable carrier. The pharmaceutical compositions are useful for inhibiting the expression of a target gene, as well as for treating diseases caused by expression of the target gene, at low dosages (i.e., less than 5 milligrams, preferably less than 25 micrograms, per kg body weight per day). The invention also relates to methods for inhibiting the expression of a target gene, as well as methods for treating diseases caused by the expression of the gene.

Nucleoside analogues expressed by the following general formula

• • where B represents an aromatic base having carbonyl oxygen at the 2-position, or a 2-hydroxyphenyl group, or oligonucleotide derivatives containing one or more of the nucleoside analogues are provided.

The oligonucleotide derivatives are triplex-forming oligonucleotide derivatives which bind specifically to target double-stranded DNA with high affinity in the antigene method to form triplexes, and can thereby control and inhibit the expression of relevant genes efficiently, and which show high resistance to nucleases.

The invention is a probe for detecting nucleic acids having a particular sequence. It is composed of two joint units. One unit is chemically different from natural nucleic acids, but has the ability to recognize a particular sequence of bases or base pairs in single or double-stranded DNA of RNA. The other unit is a compound whose detectable properties are altered upon binding to nucleic acids.

The invention relates to an approach for introducing one or more desired insertions and/or deletions of known sizes into one or more predefined locations in a nucleic acid (eg, in a cell or organism genome). They developed techniques to do this either in a sequential fashion or by inserting a discrete DNA fragment of defined size into the genome precisely in a predefined location or carrying out a discrete deletion of a defined size at a precise location. The technique is based on the observation that DNA single-stranded breaks are preferentially repaired through the HDR pathway, and this reduces the chances of indels (eg, produced by NHEJ) in the present invention and thus is more efficient than prior art techniques. The invention also provides sequential insertion and/or deletions using single- or double-stranded DNA cutting.

The present invention relates to chemically modified, linked double-stranded (ds)RNA compositions comprising two or more double-stranded (ds) oligoribonucleotides linked by at least one linking moiety and methods of formulating and delivering such compositions to modulate gene expression through target-specific RNA co-interference (RNAco-i). The compositions of the invention may optionally comprise a conjugation or a complex with one or more small molecule drugs, protein therapeutics, or other dsRNA molecules. The present invention is directed at the methods of production for, methods of use of, and therapeutic utilities for RNAi co-interference therapy utilizing the compositions of the invention.

The present invention relates to a method for inhibiting expression of a target gene, which comprises transfecting a cell, tissue, or individual organism with a double-stranded polynucleotide comprising DNA and RNA having a substantially identical nucleotide sequence with at least a partial nucleotide sequence of the target gene.

The present invention relates, e.g., to in vitro method, using isolated protein reagents, for joining two double stranded (ds) DNA molecules of interest, wherein the distal region of the first DNA molecule and the proximal region of the second DNA molecule share a region of sequence identity, comprising contacting the two DNA molecules in a reaction mixture with (a) a non-processive 5′ exonculease; (b) a single stranded DNA binding protein (SSB) which accelerates nucleic acid annealing; (c) a non strand-displacing DNA polymerase; and (d) a ligase, under conditions effective to join the two DNA molecules to form an intact double stranded DNA molecule, in which a single copy of the region of sequence identity is retained. The method allows the joining of a number of DNA fragments, in a predetermined order and orientation, without the use of restriction enzymes.