Abstract: The present invention provides a method for rapidly obtaining substantially pure DNA from a biological sample containing cells. The method involves gently lysing the membranes of the cells to yield a lysate containing genomic DNA in a high molecular weight form. The lysate is moved through a porous filter to selectively trap the high molecular weight DNA on the filter. The DNA is released from the filter using an aqueous solution to form a solution containing substantially purified DNA, from which the DNA may analyzed or recovered.

Abstract: This invention provides a rapid and highly effective method for extracting nucleic acids from cells or virions without the use of proteolytic enzymes. Extraction is accomplished within a few minutes using a lysing composition comprising a buffer, a source of a DNA polymerase cofactor, a stabilizer and at least one nonionic surfactant which will release nucleic acids from cytoplasmic and nuclear membranes of cells or virions. The resulting mixture is heated to boiling for up to fifteen minutes, and the nucleic acids are recovered for amplification using polymerase chain reaction. No proteolytic enzyme is used in the extraction process.

Abstract: An improved process for isolating A83543 factors from fermentation broth in which they are produced which comprises:a) adding an approximately equal volume of a water miscible, polar organic solvent to the fermentation broth, including the biomass thereof;b) separating the liquid phase of the resulting mixture from the biomass;c) adjusting the pH of the separated liquid phase to between about 7 and 13;d) applying the separated liquid phase directly to a column of nonfunctional, macroreticular polymer;e) eluting the A83543 components from the column with an aqueous solution of water miscible, polar organic solvent; andf) collecting the fractions containing A83543 components.

Abstract: The invention relates to a process for preparation of the vancomycin antibiotic in a microbiological way by aerobic fermentation, by using a strain belonging to the Micropolyspora species in a culture-medium containing assimilable carbon and nitrogen sources as well as mineral salts. According to the invention as microorganisms the aminoglycoside-resistant NCAIM 001092 and NCAIM 001093 mutants of the strain belonging to the Micropolyspora orientalis species are used. Nitrates are suitable mineral salts in the fermentation. Soy flour, glycerol, calcium gluconate, starch are useful carbon sources. The fermentation is carried out preferably at a temperature of 29.degree.-30.degree. C. The vancomycin antibiotic produced in the fermentation is separated by using reverse osmosis and ultrafiltration following ion-exchange chromatography.

Abstract: A methionine N.sup..alpha. -acetyltransferase enzyme has been identified in yeast. The existence of the enzyme defines a gene for methionine N.sup..alpha. -acetyltransferase which may be cloned and altered. Cells expressing the normal or variant forms of this enzyme are valuable tools for determining the amino acid sequence of uncharacterized proteins. Such cells are also valuable tools for expressing a recombinant protein lacking an acetyl group at its .alpha.-amino group.

Abstract: The invention is directed to a method of purifying sequentially synthesized peptides and oligonucleotides by affinity techniques. Selected products are capped with and N-terminus capping agent for peptides or a 5'-terminus capping agents for oligonucleotides, and then bound with affinity agents that are selective for the corresponding capping agents.

Abstract: Disclosed is a method for selecting a mutant strain belonging to the genus Streptomyces which is a hyper-producer of Streptovaricin C superior to those know heretofore. This is accomplished by first subjecting a natural strain of Streptomyces spectabilis to conditions so as to isolate organisms which are streptovaricin resistant. The streptovaricin resistant organisms thus isolated are then subjected to mutagenesis and then cultured. The colonies which are asporogenous are individually cultured in fermentation batches such that the strains take the form of pellets of varying sizes and colors. From the batch having the most heterogeneous mixture of pellets, the smallest pellet or the pellet(s) having the deepest color (usually deep red or crimson) is isolated. We have discovered that the strain of this pellet has a high likelihood of being a hyperproducer of streptovaricin. The strain of this pellet may then be subjected to fermentation conditions to produce streptovaricin.

Abstract: There is disclosed a process and a device for detecting and measuring (1) the amount of enzyme present as a detecting system following a nucleic acid hybridization reaction or immunoreaction; (2) the level and activity of free enzyme in a biological sample; (3) the level of enzyme from contaminating microorganisms present in a sample; and (4) enzymes from pure culture isolates for microbial identification and antimicrobial susceptibility testing.

Abstract: Rapid assays for analytes of interest in a fluid sample utilize a first conjugate of a labelled reactant that specifically binds to the analyte, and a second conjugate that binds to the analyte coupled to a polymer that has an affinity for a selected solid phase. The reaction components are incubated briefly, then contacted with the selected solid phase and the labelled components determined. Optional wash steps provide for enhanced sensitivity and specificity. When the analyte of interest is an antibody to HIV, the first reactant may be a synthetic, recombinant or native HIV antigen, and the second reactant may be protein A or an anti-immunoglobulin.

Abstract: The invention relates to a method of binding biologically active organic ligands to hydroxyl groups of polymeric carriers. The method involves bringing 4-fluorobenzenesulfonyl Chloride into reactive contact with the hydroxyl groups of polymeric carriers in such a manner to form sulfonate groups in place of the hydroxyl groups. The ligand is then brought into reactive contact with the hydroxyl groups of polymeric carriers to replace the sulfonate groups reacted with the organic ligand. The polymeric carrier containing the bound ligand can be used to isolate a biologically active material from a heterogeneous solution.

Abstract: A fusion protein is prepared containing a polypeptide such as an enzyme and an amino acid sequence having a substrate binding region of a polysaccharidase such as cellulase that has essentially no polysaccharidase activity. By contacting the fusion protein with an affinity matrix containing a substrate such as cellulose for the cellulase substrate binding region, the substrate binding region binds to the affinity matrix to immobilize the polypeptide. The polypeptide can be purified by separating the fusion protein or polypeptide from the affinity matrix. The polypeptide can be separated by cleaving the protein with a Cellulomonas fimi protease.

Abstract: A nucleic acid hybridization assay employing an immobilized or immobilizable polynucleotide probe selected to form DNA.RNA or RNA.RNA hybrids with the particular polynucleotide sequence to be determined. Resulting hybrids are detected by binding of an antibody reagent, preferably labeled with a detectable chemical group, selective for binding the hybrids in the presence of the single stranded sample and probe nucleic acids. No immobilization or labeling of sample nucleic acids is necessary and hybridization can be performed entirely in solution.

Abstract: Antiparasitic agents constituted by a siderophore substance including at least one hydroxamate group, at least one peptide bond, and having a molecular weight of less than 1,000 D, e.g. aerobactin. A method of isolating antiparasitic agents by purification from the supernatant of a culture of an appropriate bacteria such as pathogenic enterobacteria applicable as antiparasitic agents, in particular as antimalarial agents.

Abstract: A method for separation and recovery of polymeric beads from an antibiotic fermentation broth comprising suspending a mixture of the beads and any inherent mold from the fermentation in an aqueous solution having a specific gravity which is effective to cause the beads to form a discrete layer at or on the surface of the solution, separate and apart from the mold. The separate layer of the beads may then be easily removed from the liquid by a conventional physical methods.

Abstract: Single base pair differences between otherwise identical DNA fragments can result in an altering of the melting behavior which can be detected by denaturing gradient gelelectrophoresis (DGGE). A method has been developed for efficient transfer of genomic DNA fragments from the gel following DGGE. The DGGE is run in a polyacrylamide gel (PAG) containing 2% low gelling temperature agarose (LGT). The PAG is crosslinked with a reversible crosslinker, and after electrophoresis the crosslinks are cleaved and 80-100% of the DNA fragments are transferred to nylon membranes by alkaline transfer. Hybridization with gene specific probes is then performed. The technique has been used to identify an RFLP in the COLIA2 gene.

Abstract: The invention provides an improved process for the isolation and purification of antibiotics designated LL-E19020 alpha and beta which are derived from the microorganism Streptomyces lydicus ssp. tanzanius NRRL 18036.The invention is a method of recovery, concentration from crude solutions and to processes for the purification of anitbiotics designated LL-E19020 alpha and beta which are derived from the microorganism Streptomyces lydicus ssp. tanzanius NRRL 18036.

Abstract: The present invention provides a method for rapidly obtaining substantially pure DNA from a biological sample containing cells. The method involves gently lysing the membranes of the cells to yield a lysate containing genomic DNA in a high molecular weight form. The lysate is moved through a porous filter to selectively trap the high molecular weight DNA on the filter. The DNA is released from the filter using an aqueous solution to form a solution containing substantially purified DNA, from which the DNA is recovered.

Abstract: A method of removing an organic solvent from a mixture that includes a compound having a polypeptide segment and the organic solvent, the method including the steps of contacting the mixture with an ion exchange resin under conditions that allow the compound to bond to the resin; and washing the resin with a first aqueous solution that elutes the organic solvent from the resin while allowing the compound to remain bound to the resin.

Abstract: The invention describes a method for extracting DNA from a sample. It involves contacting a sample with a separation reagent which permits differentiated solvation of DNA and protein. By adding a gel polymer, such as a silical gel polymer to the mixture of sample and separating reagent followed by agitation via, e.g., centrifugation, the DNA and protein are separated, with the gel acting as a block to prevent contamination of the DNA phase by the protein. Higher yields of DNA are obtained as compared to methodologies where the gel is not used. Additionally, the problems associated with the physical contact of the solvents, which are frequently carcinogens, are avoided. Also taught are kits which can be used in connection with the inventive method.

Abstract: Purified endotoxin protein is recovered from aqueous suspensions containing lysed cells and Bacillus thuringiensis crystalline endotoxin protein in a multistep process.The aqueous suspension, e.g., a concentrated fermentation culture, is first treated under strongly basic or acidic conditions to solubilize the crystal protein and the solution is then separated from residual solids. Endotoxin protein is precipitated from the separated aqueous solution, by adjusting its pH to the isoelectric point of the protein, and recovered by centrifugation or the like.

Abstract: A method is provided for removing an anion from an aqueous liquid, such as a fermentation broth. The aqueous broth is contacted with a water-immiscible ion exchange liquid to extract the anion from the broth. The anion exchange liquid is then back extracted with an aqueous phase, to remove the anion, preferably for other uses.

Abstract: A method for separation and recovery of polymeric beads from an antibiotic fermentation broth comprising suspending a mixture of the beads and any inherent mold from the fermentation in an aqueous solution having a specific gravity which is effective to cause the beads to form a discrete layer at or on the surface of the solution, separate and apart from the mold. The separate layer of the beads may then be easily removed from the liquid by a conventional physical methods.

Abstract: A process is provided for isolating and purifying biologically active RNA from a biological sources containing RNA, DNA and other cellular materials. The process involves contacting the RNA-containing source with particles comprising siliceous material, such as finely-divided glass, in the presence of a binding solution comprising concentrated, acidified chaotropic salt. Under these conditions, RNA, but not DNA, binds selectively to the siliceous material, and can be separated easily from the other components of the sample. Preferably, the selective binding process is applied to biological cells containing RNA of interest. Intact cells are disrupted by exposing them to a lysing solution comprising, as its main component, concentrated, acidified chaotropic salt. The RNA is then isolated and purified from the lysate using the selective binding process of the invention.

Abstract: Basement membrane proteins are isolated as functioning proteins in relatively large amounts from human or animal tissues in aqueous solution in the presence of a chelating agent. It is possible to use these proteins to obtain highly specific antibodies which are used for the immunological determination of these proteins.

Abstract: A fusion protein is prepared containing a polypeptide such as an enzyme and an amino acid sequence having a substrate binding region of a polysaccharidase such as cellulase that has essentially no polysaccharidase activity. By contacting the fusion protein with an affinity matrix containing a substrate such as cellulose for the cellulase substrate binding region, the substrate binding region binds to the affinity matrix to immobilize the polypeptide. The polypeptide can be purified by separating the fusion protein or polypeptide from the affinity matrix.

Abstract: A process is disclosed for chemically converting a substrate into its reaction products and immediately thereafter physically separating the reaction products in a continuous operation. The process is carried out with a bioreactor having an ultrafiltration membrane containing an immobilized chemical agent which is preferably an enzyme. The bioreactor is prepared by securing an ultrafiltration membrane to an inside wall of a porous tubular support and chemically bonding an enzyme to an inner surface of the membrane. The enzyme is preferably bonded to the membrane by chelation and the membrane may be a polymeric membrane or a metal oxide membrane. To convert a substrate, a substrate-containing feed stream is preferably flowed tangentially along the inner surface of the membrane containing the immobilized enzyme. Sufficiently small reaction products filter through pores of the membrane and larger reaction products are retained by the membrane.

Abstract: The lactosucrose high-content powder according to the present invention is prepared by allowing an aqueous solution containing sucrose and lactose to act on a saccharide-transferring enzyme, removing concomitant saccharides in the resultant saccharide solution containing lactosucrose to obtain a lactosucrose high-content solution with a lactosucrose content of 45 w/w % or higher on sugar composition, and spray-drying the resultant solution to obtain a lactosucrose high-content powder. The powder is incorporated in an orally- or parenterally-administrable product to obtain an orally- or parenterally-administrable product which exerts a selective growth-promoting-effect on a microorganism of the genus Bifidobacterium in vivo.

Abstract: In a process for growing enzyme crystals, small crystals are continuously removed from a crystallizer, dissolved and returned to the crystallizer to maintain a supersaturated state. The method permits the growing of large crystalline enzymes of uniform size of about 0.5 to 1 mm. Solid materials can be coated with crystalline enzymes by placing a solid material in the crystallizer such that crystals deposit on the solid material. The process is preferably used to produce crystalline glucose isomerase.

Abstract: Novel methods for assaying a nucleic acid analyte are provided, which employ polynucleotides having oligonucleotide sequences substantially homologous to a sequence of interest in the analyte, where the presence or absence of hybridization at a predetermined stringency provides for the release of a label from a support. Particularly, various techniques are employed for binding a label to a support, whereupon cleavage of either a single or double strand, a label may be released from a support, where the release of the label can be detected as indicative of the presence of a particular oligonucleotide sequence in a sample. The method finds use in diagnosis of disease, genetic monitoring, and analysis of nucleic acid mixtures.

Abstract: The invention relates to a process for the preparation of indole applicable in flavoring and perfume compositions wherein a micro-organism which does not or hardly metabolize indole is cultured aerobically or anaerobically in a culture medium containing as the substrate tryptophan of natural origin. The produced indole in the fermentation broth may be isolated therefrom with a food grade extraction agent.

Abstract: A method is provided for controlling production of low molecular weight heparin (LMW-heparin) when depolymerizing heparin with heparinase in a reaction mixture in a reactor. Depolymerization to a desired average molecular weight is monitored by measuring an increase in UV-absorption (preferably at 230-235). When the absorption has reached a value for a desired molecular weight, the depolymerization is stopped or LMW-heparin having the desired molecular weight is continuously removed from the reaction mixture. In a preferred embodiment, the reaction mixture is subjected to ultrafiltration to produce a filtrate containing LMW-heparin, and a retentate which is recycled to the reactor. UV-absorption and refractive index of the filtrate are measured, and depolymerization is controlled in accordance with the measured absorption and refractive index to produce a filtrate containing a LMW-heparin of low polydispersity and predetermined molecular weight.

Abstract: The invention describes a method for extracting DNA from a sample. It involves contacting a sample with a separation reagent which permits differentiated solvation of DNA and protein. By adding a gel polymer, such as a polyester gel polymer to the mixture of sample and separating reagent followed by agitation via, e.g., centrifugation, the DNA and protein are separated, with the gel acting as a block to prevent contamination of the DNA phase by the protein. Higher yields of DNA are obtained as compared to methodologies where the gel is not used. Additionally, the problems associated with the physical contact of the solvents, which are frequently carcinogens, are avoided. Also taught are kits which can be used in connection with the inventive method.

Abstract: A process for the fermentative production of the deoxyribonucleoside thymidine and/or its corresponding base thymine by aerobically cultivating a strain of the genus Brevibacterium, in particular one of the strains NCIMB 40117 and 40116. The produced thymidine may be used as an intermediate in the production of azidothymidine and active ingredient in a composition for use in the treatment of auto imune deficiency syndrome (AIDS). Biologically pure cultures of strain NCIMB 40014 and variants and mutants derived therefrom are claimed per se.

Abstract: A novel urease having an optimal pH for activity in the acidic region is produced by a microorganism belonging to the genus Lactobacillus or Streptococcus. The urease is superior to the conventional urease in pH stability, temperature stability and alcohol stability.

Abstract: In a method of the separation of yeasts from a fermentation liquor contained in a fermentation tank the fermentation liquor is forced by a pump through a conduit system into a filter container in which the fermentation liquor is forced through a number of vertically suspended candle-like filters so that yeast filter cakes are formed on the filter cloth with water of the filters. The cakes are then washed with water, blown off with pressurized air for dewatering, loosened with a counterflow of a pressurized gas and discharged from the filter container. The filter container is immediately connected to the fermentation tank.

Abstract: A method is disclosed for the production of xylitol from a xylose and/or xylan containing material. A solution containing xylitol and other hexoses is fermented with a yeast strain capable of converting free xylose to xylitol and other free hexoses present to ethanol, with the xylitol subsequently separated by chromatographic separation means.

Abstract: A solid support useful for bioaffinity and ion-exchange separations and enzyme immobilization is provided. The support is based on a non-perfluorocarbon solid carrier core coated with a nonionic fluorosurfactant-coated fluorocarbon interlayer to which a ligand or a binder for the ligand is securely, but reversibly attached through a reactive perfluorocarbon anchor compound. Also provided is a solid support useful for size exclusion separations. Such support is based on a non-perfluorocarbon carrier core coated with a nonionic fluorosurfactant-coated fluorocarbon interlayer.

Abstract: This invention is directed to a process for the purification of plasmid and other DNA, both single-stranded and double-stranded, by immobilizing the DNA onto diatomaceous earth in the presence of a chaotropic agent and eluting the DNA with water or low salt buffer. The resulting purified DNA is biologically active. Also included in the invention is a process for the immobilization of DNA onto diatomaceous earth in the presence of a chaotropic agent.

Abstract: A DNA probe has been isolated which is capable of hybridizing to an oligonucleotide sequence coding for a polypeptide from a major 64 Kilodalton protein of human cytomegalovirus (HCMVgp64). The probe has a sequence of at least seventeen (17) to as many as seven hundred twenty-one 721) nucleotides. The probe may be labelled as by radioactivity. The probe has been used to screen DNA fragments constituting a subgenomic library of human cytomegalovirus DNA to obtain DNA fragments coding for the major late protein of human cytomegalovirus. The DNA fragments coding for the major late protein of human cytomegalovirus (HCMVgp64) may be hybridized to DNA fragments of HCMV DNA from an individual having human cytomegalovirus infection. The viral DNA can be used as whole HCMV DNA or as fragments formed by digesting the human cytomegalovirus DNA with a restriction endonuclease such as one of the restriction endonucleases EcoRI, BamHI, XbaI, HindIII and PrtI.

Abstract: The invention relates to a process for the in vitro detection of a determined RNA, particularly of a mRNA connected with a genetic abnormality in a biological material. This process comprises a treatment of the cells contained in that material for the sake of releasing their cellular components and of exposing the RNAs yet without denaturing other nucleic acids, and then the detection operation comprising contacting RNA sought to be detected, in presence of the other cellular components, with a nucleotidic sequence complementary of the RNA sought.

Abstract: Specific sialyl transferases (ST) are important enzymes in the manufacture of carbohydrate moieties useful for pharmacological purposes and for diagnostic and clinical procedures. The invention offers simplified methods to isolate highly specific forms of these ST enzymes by taking advantage of the affinity of the ST for its acceptor substrate in the presence of the sialyl-CMP analogs, such as CDP. Specifically exemplified are isolation of the alpha 2,3-ST, specific for Le.sup.c or LacNAc, and alpha 2,6-ST specific for LacNAc.

Abstract: Peptide preparation is carried out by a continuous process by supplying a serine protease or peptidase enzyme retained in a reaction vessel with an alkyl ester of an N-protected amino acid or oligopeptide and a recycle stream containing an amino acid or oligopeptide to form an N-protected chain extended peptide, separating the N-protected chain extended peptide by adsorption on a hydrophobic absorbent, and eluting and recovering the adsorbed N-protected chain extended peptide. Adsorption is carried out without adjusting the pH from that in the reaction vessel, and adsorber effluent is recycled to the recycle stream. The protease or peptidase enzyme may be immobilized, and there is substantial exclusion of organic solubilizers in the reaction vessel. A preferred N-protecting group is an N-phenacyl group. The N-protecting group can be separated from the N-protected chain extended peptide with a deprotecting enzyme to recover the chain extended peptide.

Abstract: A composition for removing metal ions from aqueous solution is prepared by immobilizing metal ion-binding microorganisms such as algae, washing the immobilized microorganisms, drying the washed immobilized microorganisms and heating the dried immobilized microorganisms to a temperature of about 300.degree. to about 500.degree. C. for a time sufficient to provide a stable composition that is non-swelling in aqueous solution. The composition preferentially adsorbs precious metal ions from an aqueous solution containing concentrations of base metal ions and/or other dissolved materials several orders of magnitude greater than the concentration of the precious metal ions. The composition can also be used to extract precious metal ions from geothermal fluids.

Abstract: In order to carry out hybridization reactions by forming a complex between a polynucleotide and a detectable substance and using the complex thus formed for hybridization reactions, a single strand polynucleotide is reacted with a positively charged nucleic acid binding detectable protein or polyamine in solution and the resultant complex is covalently bound with a cross-linking agent and the polynucleotide compound obtained is used as a marker in hybridization experiments in the elucidation of complementary sequences in foreign polynucleotides. The protein may be an enzyme such as horseradish peroxidase modified by a polyamine e.g. a polyethylene imine to provide a positively charged region. Assay sensitivity may be enhanced by performing the hybridization in the presence of about 6% (w/v) of a polyethylene glycol.

Abstract: Nucleic acid encoding lecithin:cholesterol acyltransferase is ligated into expression vectors and used to transform host cells for the synthesis of lecithin:cholesterol acyltransferase in recombinant cell culture. Lecithin:cholesterol acyltransferase amino acid sequence variants are described for enhancing the properties of lecithin:cholesterol acyltransferase. Lecithin:cholesterol acyltransferase and its variants are employed in the therapy of conditions characterized by hypercholesterolemia and for the mobilization of cholesterol in vivo.

Abstract: A composition for isolating and purifying nucleic acid from cell culture medium and a method for isolating and purifying nucleic acid from cell culture medium employing the composition, in which the reagent includes about 1 to 3.5M acetate salt solution, about 4 to 11.2M acetic acid, about 1 to 40% by volume phenol, and about 1 to 40% by volume chloroform.

Abstract: A DNA probe has been isolated which is capable of hybridizing to an oligonucleotide sequence coding for a polypeptide from a major 64 Kilodalton protein of human cytomegalovirus (HCMVgp64). The probe has a sequence of at least seventeen (17) to as many as seven hundred twenty-one (721) nucleotides. The probe may be labelled as by radioactivity. The probe has been used to screen DNA fragments constituting a subgenomic library of human cytomegalovirus DNA to obtain DNA fragments coding for the major late protein of human cytomegalovirus. The DNA fragments coding for the major late protein of human cytomegalovirus (HCMVgp64) may be hybridized to DNA fragments of HCMV DNA from an individual having human cytomegalovirus infection. The viral DNA can be used as whole HCMV DNA or as fragments formed by digesting the human cytomegalovirus DNA with a restriction endonuclease such as one of the restriction endonucleases EcoRI, BamHI, XbaI, HindIII and PrtI.

Abstract: A process for removing endotoxin from Gram-negative polysaccharides such as H. influenzae polyribosylribitol phosphate by adding alcohol incrementally until substantially all lipopolysaccharide precipitates.

Abstract: Crystalline subtilisin is produced by adding a halide salt, such as sodium chloride or calcium chloride, to a concentrated subtilisin solution (at least about 40 g/1). This process does not produce amorphous subtilisin even at high salt concentrations in the solution. Optionally, subtilisin seed crystals also may be added to the concentrate to speed up the crystallization process.

Abstract: A process for removing endotoxin from Gram-negative polysaccharides such as polyribosylribitol phosphate by solubilizing polysaccharide-containing powder derived from Gram-negative bacteria fermentation broth to provide a divalent counter ion for endotoxin and adding alcohol incrementally to induce lipopolysaccharide precipition, and mixing material resulting from the alcohol addition with a nonionic resin, a detergent and a chelating agent.