Abstract: The present invention is a general method for irreversibly inactivating ribonucleases. Ribonucleases are completely inactivated by treating them with a reducing agent and heat. RNA samples contaminated with ribonuclease may be treated with this method to protect them from degradation. The RNA may then be used directly in a variety of enzymatic reactions and molecular biology techniques. This method may also be applied to a variety of molecular biology reagents which may be contaminated with ribonuclease to protect an RNA from being degraded when incubated with the reagent.

Abstract: The present invention relates to methods for pretreating biological samples for extraction of nucleic acid therefrom The present invention employs a combination of at least one protein denaturant with one or more of the following elements to form a reaction mixture for extraction of nucleic acid: (1) at least one aprotic solvent, (2) stepwise heating, and (3) sample dilution.

Abstract: A method for isolating RNA, DNA or proteins from an animal tissue sample that is amenable to high throughput adaptation is disclosed. The method involves beadmilling the sample to disrupt the cells contained therein and extracting RNA, DNA or proteins from the disrupted sample by solid phase extraction.

Abstract: The invention relates to a method for preparing biological samples for analysis, comprising the following steps: (a) the biological sample is placed on a two-dimensional support; (b) application of a protein-precipitating or denaturing first solution L1 to the biological sample at a first temperature T1 for a predetermined first time period Z1; (c) the protein-precipitating or denaturing first solution L1 is then left, or more solution is applied to the biological sample or a protein-precipitating or denaturing second solution L2 is applied to the biological sample at a temperature T2, for a predetermined second time period Z2, whereby T2 is lower than T1 and Z2 is longer, equal to or shorter than Z1; and (d) drying of the sample.

Abstract: The invention describes a method for the isolation of components from samples, particularly large molecular weight DNA from biological samples. The method involves the application of controlled oscillatory mechanical energy to the sample for short periods of time of about 5 to 60 seconds to lyse the sample and release the component(s) from the sample, followed by standard isolation methods. In preferred embodiments, the method includes the use of a spherical particle for applying the mechanical energy.

Abstract: A process for the recovery of plasmids or other DNA from cells using a first filtration step to remove the cellular debris and other large cellular components and then an ultrafiltration step to capture the plasmids or other DNA on the surface of the ultrafiltration membrane where they may be recovered. An apparatus is also taught for enacting the process and comprises an upper microfiltration or coarse filtration membrane and a lower ultrafiltration membrane. The driving force may be the same for both filters or different and may be done sequentially or simultaneously.

Abstract: Methods and compositions are provided for extracting DNA from tissues, hair, teeth, bone, plant material, solid matrices and other samples from which DNA extraction is generally regarded as being difficult. DNA may quickly be extracted from samples, for example, in just 5 minutes to 2 hours, without enzymatic digestion or the use of toxic organic chemicals such as phenol, chloroform, guanidine thiocyanate or 2-mercaptoethanol.

Abstract: Methods and compositions for extracting nucleic acids from a biological sample are provided. The extraction compositions contain a protease enzyme such as proteinase K at alkaline pH with little or no surfactant present. Extraction can be efficiently performed in 60 minutes or less at room temperature for certain mammalian tissue samples and at elevated temperatures for certain plant tissues.

Abstract: Method for processing a biological sample contained in a liquid, by

Abstract: Cell lysis compositions, methods for extracting and isolating proteins and peptides from a host cells using the compositions, kits and apparatus for extracting and isolating protein and peptide molecules from host cells and for detecting for the presence of a protein or peptide. The composition allows for the extraction and isolation of proteins and peptides from host cells without the need for mechanical disruption and with or without isolation of the cells from cell medium. The composition includes at least one surfactant having a hydrophobic-lipophilic balance value in the range from about 11 to about 16; and at least one cell membrane altering compound.

Abstract: Methods are disclosed for rapid, reliable and simple isolation of RNA, DNA and proteins from formalin-fixed paraffin-embedded tissue samples. RNA purified in this manner can be used to monitor gene expression levels. The tissue sample can be a tumor or other pathological tissue.

Abstract: This invention describes a rapid (10 to 15 minutes), simple, flexible and efficient method of nucleic acids extraction for nucleic acid testing assays. This method has the following basic steps: i) mechanical cell lysis using solid particles in the presence of a chelating agent, followed by ii) controlling the presence and/or activity of NAT assays inhibitors. This method is applicable to various biological samples and universal for microorganisms, as one can use it to extract nucleic acids from test samples containing target viruses, bacteria, bacterial spores, fungi, parasites or other eukaryotic cells, including animal and human cells.

Abstract: The present invention provides a method and a system for the co-isolation of cognate DNA, RNA and protein sequences, a method for screening co-isolates for defined activities, and the production of operons for simultaneous or coordinated expression of genes. Encapsulating solutions and microstructures in lipid vesicles or polymer capsules permits transcription/translation reactions amenable to techniques and instruments that utilize an aqueous environment. Microstructures containing binding sites for both mRNA and protein and encapsulated into a liposome or hollow polymer capsule, creating a reactosome. After transcription/translation, or other reactions, the reactosomes may be screened using aqueous methods. Once the desired reactosomes are isolated, the vesicles are disrupted, and the microspheres can be collected. Once transcription/translation reactions occur, the capsules may be frozen for storage or later use.

Abstract: The invention relates to a method of separating extra-chromosomal DNA from RNA. It also relates to DNA produced by the method and pharmaceuticals derived from such DNA, for example, DNA vaccines.

Abstract: The present invention provides a method of extracting virus genes, which is excellent in quickness and convenience and can be automated, and this invention also provides an apparatus for such extraction of virus genes. A conductive hollow fiber membrane (module) 12 is immersed in a virus suspension 20 and thus viral particles suspended in the virus suspension 20 are captured in the conductive hollow fiber membrane (module) 12. The coating of the thus captured virus is lysed and the virus genes are released. Next, the virus genes are collected by subjecting the conductive hollow fiber membrane (module) 12 to at least one of an electrical treatment and a physical treatment.

Abstract: The present invention provides a method of isolating nucleic acid from a sample of cells, said method comprising: (a) binding cells in said sample in a solid support to isolate cells from the sample; (b) lysing the isolated cells; and (c) binding nucleic acid released from said lysed cells to said same solid support and a kit for carrying out such a method. The method may advantageously be used to prepare nucleic acid for use in a nucleic acid-based target cell detection method.

Abstract: A process is disclosed for the extraction and isolation of the extrachromosomal DNA from blood and cells with a view to detecting DNA alterations linked to a possible pathology in progress and making a diagnosis. The process for DNA extraction comprises the use of a buffer, a heat treatment, followed by treatment with a detergent and a salt. The extract thus obtained is suitable for undergoing electrphoresis on agarose gel.

Abstract: An object of the present invention is to provide a method for separating and purifying a nucleic acid by adsorbing the nucleic acid in a test sample to a surface of a membrane and desorbing the nucleic acid by washing and the like.

Abstract: The invention describes a method for the isolation of components from samples, particularly large molecular weight DNA from biological samples. The method involves the application of controlled oscillatory mechanical energy to the sample for short periods of time of about 5 to 60 seconds to lyse the sample and release the component(s) from the sample, followed by standard isolation methods. In preferred embodiments, the method includes the use of a spherical particle for applying the mechanical energy.

Abstract: Reagents and method for genotyping mice and other animals by in situ Polymerase Chain Reaction amplification of a target polynucleotide in the tissue biopsy. The reagent is comprised of non-ionic detergents, a protease, a buffering agent, a metal ion cofactor, a chelating agent and a salt. The method is comprised of taking a tissue biopsy; admixing it with the reagent; a Lysing Cycle, an inactivation cycle, an amplification step and a detection step.

Abstract: A method for purifying nucleic acids wherein objective nucleic acids are separated through solid-liquid separation by deposition of the nucleic acids onto a solid-phase carrier, via enzymatic treatment of a cell lysis solution, from a sample containing nucleated cells and the like, as well as a kit for carrying out the method.

Abstract: A method of cleaning a pin head or other implements between handling a plurality of samples of nucleic acid subject to an amplification process using a biochemical amplification product. The method comprises exposing the pin head or other implement to ultraviolet (UV) and preferably also infrared (IR) illumination between handling different samples in order to suppress cross-contamination by non-specific amplification. The biochemical amplification product may be obtained from a rolling circle amplification (RCA) procedure, such as that catalyzed by bacteriophage phi29, or a polymerase chain reaction (PCR) amplification product.

Abstract: Methods are disclosed for rapid, reliable and simple isolation of RNA from formalin-fixed paraffin-embedded tissue samples. RNA purified in this manner can be used for quantitative measurement gene expression levels. The tissue sample can be a tumor or other pathological tissue.

Abstract: The invention relates to a method for the integrated treatment of biomass from a cell culture process for producing clear cell lysate containing plasmid DNA. According to the invention, in a first step filtering occurs in a filter in the presence of a filtering agent. In a second step, the biomass contained in the filter cake which is obtained is thermally decomposed and the cell lysate is filtered in another step. The obtained cell lysate containing plasmid DNA is characterised by a clarity of OD600 of at a maximum of 0.05 E/cm and which is suitable for cloning, transformation, transfection, microinjection in cells, gene therapy, DNA vaccination and/or for polymerase chain reaction (PCR).

Abstract: This invention relates to the discovery that both non-particle and particle associated nucleic acids are present in blood plasma and serum and can be used to evaluate disease conditions.

Abstract: Methods and compositions are provided for the isolation of genomic DNA (gDNA) and Bacterial Artificial Chromosomes (BACS) from a target source. Novel nucleic acid trapping membranes are also provided, the membranes composed of oleophobic coated or treated glass and/or acrylic fibers or beads.

Abstract: The invention is in the area of selective extraction of DNA from groups of cells. Selective lysis of a particular cell type within a cellular mixture is performed and then the mixture is separated with a filter that allows the DNA from the lysed cells to flow through the filter, while not allowing the unlysed cells to pass through, thereby selectively extracting the DNA from a particular cell type. In one specific embodiment, spermatozoa DNA can be isolated from biological samples which also contain epithelial cells. Methods and kits are also provided which allow for the sequential extraction of DNA from mixtures of cells. The DNA in the sample can be from human, animal or vegetal origin, or any combination of human, animal or vegetal DNA.

Abstract: A process for the recovery of plasmids or other DNA from cells using a first filtration step to remove the cellular debris and other large cellular components and then an ultrafiltration step to capture the plasmids or other DNA on the surface of the ultrafiltration membrane where they may be recovered. An apparatus is also taught for enacting the process and comprises an upper microfiltration or coarse filtration membrane and a lower ultrafiltration membrane. The driving force may be the same for both filters or different and may be done sequentially or simultaneously.

Abstract: The invention concerns a process for preparing biological samples for the subsequent detection of an analyte. In particular, the invention relates to a process for the isolation of a nucleic acid in a sample using a suspension of magnetic glass particles. In addition, kits and apparatuses containing magnetic glass particles for sample preparation are provided.

Abstract: A method for isolating a polynucleotide of interest that is present in the genome of a first mycobacterium strain and/or is expressed by the first mycobacterium strain, where the polynucleotide of interest is also absent or altered in the genome of a second mycobacterium strain and/or is not expressed in the second mycobacterium. The method includes (a) contacting the genomic DNA of the first mycobacterium strain under hybridizing conditions with the DNA of a least one clone that belongs to a bacterial artificial chromosome (BAC) genomic DNA library of the second mycobacterium strain, and (b) isolating the polynucleotide of interest that does not form a hybrid with the DNA of the second mycobacterium strain. This invention further pertains to a Mycobacterium tuberculosis strain H37Rv genomic DNA library, as well as a Mycobacterium bovis BCG strain Pasteur genomic DNA library, and the recombinant BAC vectors that belong to those genomic DNA libraries.

Abstract: This invention provides methods of obtaining novel polynucleotides and encoded polypeptides by the use of non-stochastic methods of directed evolution (DirectEvolution™). A particular advantage of end-selection-based methods is the ability to recover full-length polynucleotides from a library of progeny molecules generated by mutagenesis methods. These methods include non-stochastic polynucleotide site-saturation mutagenesis (Gene Site Saturation Mutagenesis™) and non-stochastic polynucleotide reassembly (GeneReassembly™). This invention provides methods of obtaining novel enzymes that have optimized physical &/or biological properties. Through use of the claimed methods, genetic vaccines, enzymes, small molecules, and other desirable molecules can be evolved towards desirable properties. For example, vaccine vectors can be obtained that exhibit increased efficacy for use as genetic vaccines.

Abstract: The present invention relates to a method of isolating nucleic acid from a blood sample, said method comprising: (a) selectively isolating leucocytes from said sample by binding said leucocytes to a solid support by means of a binding partner specific for leucocytes; (b) lysing said isolated leucocytes; and (c) binding nucleic acid released from said lysed cells to said solid support. Kits for isolating nucleic acid from samples form further embodiments of the invention.

Abstract: This invention concerns a method for the lysis of prokaryotic or eukaryotic cells, or for simultaneous lysis of both prokaryotic and eukaryotic cells

Abstract: The invention relates to methods for the detection of a specific sequence of RNA in a cell or tissue sample. The invention also relates to methods to enzymatically manipulate the RNA in a crude cell lysate in a number of applications.

Abstract: For reducing side effects of blood transfusion, white (nucleated) blood cells are separated from donated human blood by filtering the blood through filter media selectively adsorbing white blood cells. Such filter media are processed for recovering nucleic acids contained in the white blood cells. The cells retained by the filter media are separated from the filter media e.g. by washing with an aqueous solution. The separated cells are then lysed and the nucleic acids are isolated from the components of the lysed cells. Preferably each portion of filter medium is used for filtering the blood of one individual donor and is processed separately whereby samples of nucleic acids of one individual each are obtained.

Abstract: The present invention provides a method of isolating nucleic acid from a sample of cells, said method comprising: (a) binding cells in said sample to a solid support to isolate cells from the sample; (b) lysing the isolated cells; and (c) binding nucleic acid released from said lysed cells to said same solid support and a kit for carrying out such a method. The method may advantageously be used to prepare nucleic acid for use in a nucleic acid-based target cell detection method.

Abstract: The present invention relates to the use of compositions for isolating and/or stabilising nucleic acids in or from microorganisms—such as prokaryotes, fungi, protozoa or algae.

Abstract: An improved method for isolating DNA from biological samples is provided.

Abstract: The entire process of reverse transcription-polymerase chain reaction (RT-PCR) is simplified by using oligonucleotide-immobilized microplates made of, e.g., polypropylene, to which oligonucleotides are securely immobilized and which can be subjected to thermal cycles of PCR. RT-PCR is preferably conducted in solid-phase. Capturing of mRNA and RT-PCR can be conducted in the same plates. The cDNA synthesized from the mRNA captured on the microplates can be used more than once. Further, in combination with the microplates, a filter plate is used for the preparation of cell lysates wherein target cells are placed on the filter plate, and a lysis buffer is passed through the cell layer on the filter to transfer cell lysate directly to the microplate via well-to-well communication.

Abstract: Disclosed are compositions and a method for amplification of nucleic acid sequences of interest. The disclosed method generally involves replication of a target sequence such that, during replication, the replicated strands are displaced from the target sequence by strand displacement replication of another replicated strand. In one form of the disclosed method, the target sample is not subjected to denaturing conditions. It was discovered that the target nucleic acids, genomic DNA, for example, need not be denatured for efficient multiple displacement amplification. The primers used can be hexamer primers. The primers can also each contain at least one modified nucleotide such that the primers are nuclease resistant. The primers can also each contain at least one modified nucleotide such that the melting temperature of the primer is altered relative to a primer of the same sequence without the modified nucleotide(s). The DNA polymerase can be &phgr;29 DNA polymerase.

Abstract: The invention relates to a method for isolating plasmids from suspended bacterial or yeast cells using a filter matrix, at least one lysing substance being added to the suspension and pre-damaging or completely lysing the predominant portion of the suspended cells, at least one conformation-altering substance being added to said suspension to alter the conformation of the plasmids to be isolated in such manner as to promote retention of the cell components in or at the filter matrix, the suspension thereupon being moved through a filter matrix and the cells if not yet lysed then being lysed to completion upon contact with the filter matrix and the released plasmids being retained in or at the said filter matrix.

Abstract: Disclosed is a method of extracting nucleic acids from blood which comprises contacting blood cells, preferably after lysing with an activated solid phase at one pH to immobilize the nucleic acids and then removing the nucleic acids at a higher pH when the charge has been reversed or neutralized. The solid phase can be beads activated by a histidine as a binding agent. The beads can be fluidized by sucking the blood with air up through a column containing the beads to improve contact and prevent clogging.

Abstract: A method for preparing a nucleic acid component of a sample for amplification includes contacting the sample with a porous support that deactivates a nucleic acid amplification inhibitor component of the sample and directing a fluid through the porous support, whereby the nucleic acid component of the sample is directed through at least a portion of the porous support and is separated from the support, thereby preparing the nucleic acid component for amplification.

Abstract: Methods and compositions for the identification, isolation and characterization of regulatory DNA sequences in a cell of interest are provided. Also provided are libraries of regulatory sequences obtained according to the methods, and databases comprising collections of regulatory sequences for a particular cell of interest. In addition, various uses for the regulatory sequences so obtained, and uses for the databases of regulatory sequences, are provided. Also disclosed are computer systems and computer program products for utilizing the databases to conduct various genetic analyses, and uses of accessible regulatory sequences in the design of vectors bearing transgenes.

Abstract: In one embodiment, methods are provided for differential isolation of cellular contents from different cells for further analysis. In one exemplary embodiment, the nucleic acids of bacteria are isolated from a mixture of bacteria and animal cells by lysing animal cells.

Abstract: It is one objective of the present invention to obtain reproducible representations of expressed mRNA molecules by exploiting a novel technique that relies on short, single stranded polynucleotide tags. In one preferred embodiment, only one polynucleotide tag is obtained from each mRNA molecule, and relatively simple counting statistics can thus be applied after identification and sampling of the different tags, or a subset of tags being present in the population of representative tags. The tags according to the present invention are preferably single stranded polynucleotide tags obtained by subjecting genetic material derived from a biological sample to at least one site-specific nicking endonuclease capable of i) recognizing a predetermined nucleotide motif comprising complementary nucleotide strands and ii) cleaving only one of said complementary strands in the process of generating the at least one single stranded polynucleotide tag.

Abstract: An object of the present invention is to provide a novel method for suppressing the action of nucleic acid synthesis inhibitory substances and thereby amplifying a nucleic acid in a sample efficiently. According to the present invention, in a method for synthesis of nucleic acids to amplify an intended nucleic acid in a sample, the sample is brought in advance into contact with an insoluble polymer of a polyanion, a sulfated polymer or a sulfated polysaccharide, to remove nucleic acid synthesis inhibitory substances.

Abstract: The present invention includes a method for detecting and isolating sugarcane proteins that interact with the HC-Pro and P1 proteins of SrMV and other proteins involved in gene silencing, particularly in sugarcane. The method uses a two hybrid assay with an HC-Pro, P1, or other silencing-related protein-containing bait protein and a prey protein containing a polypeptide encoded by a DNA molecule in a cDNA library. The method also includes identification of false positives through reverse two-hybrid assays and using in vitro techniques such as farwestern blots or pull down assays where plant physiological conditions may be replicated. Finally, interactions may be confirmed in planta. Some novel proteins used in and discovered using the these methods are also identified. Methods of using viral and plant proteins to regulate silencing in plants such as sugarcane are also discussed.

Abstract: The present invention relates to a method of purifying a set of specific DNA molecules to be used in DNA-DNA hybridisations, as well as to DNA probes containing less than 2% Cot-1 DNA.

Abstract: A method for manipulating genetic material, the method comprising disrupting cells so as to liberate genetic material contained in the cells; contacting the genetic material to a silica column in a manner to cause the genetic material to become immobilized to the column; labeling the immobilized genetic material; and eluting the labeled material from the column. Also provided is a two-buffer process for manipulating genetic material, the process comprising: contacting cells containing the genetic material to a silica column; creating a first fraction of cell detritus and a second fraction containing the genetic material; confining the genetic material to the column; removing the cell detritus; contacting the genetic material with radicals so as to produce reactive aldehyde groups on the genetic material, and attaching chromophore to the genetic material.