Abstract: In some embodiments the present disclosure provides a composition for targeted alteration of a DNA sequence and methods of altering the targeted DNA sequence using the composition. In some embodiments such a composition comprises a MiniVector comprising a nucleic acid sequence template for homology-directed repair, alteration, or replacement of the targeted DNA sequence within a cell in vivo or in vitro, where the MiniVector lacks both a bacterial origin of replication and an antibiotic selection gene, and wherein the Mini Vector has a size up to about 2,500 base pairs.

Abstract: The invention provides compositions and methods for reprogramming minimal volumes of mononuclear cells. In particular aspects, the invention provides methods and compositions for reprogramming minimal volumes of umbilical cord blood obtained from cord blood segments from cryopreserved cord blood segments.

Abstract: The invention relates to vectors and mammalian cells in a system useful for switching on or switching off gene expression in response to pH and gaseous carbon dioxide, taking advantage of signal transduction of the human proton-activated cell-surface receptor TDAG8, OGR1 and/or GPR4 and related receptors to chimeric promoters containing cAMP-response elements. The systems according to the invention can be used to precisely monitor culture pH within the physiologic range, or also to adjust expression by gaseous C02 which shifts the C02-bicarbonate balance towards hydrogen ions. Production of biopharmaceuticals of engineered cells cultivated in a bioreactor setup can be induced by gaseous C02. Also, implanting engineered cells provide novel treatment strategies for acidosis-related disorders and type 1 diabetes.

Abstract: The invention relates to compositions and methods for the preparation, manufacture and therapeutic use of polynucleotides, primary transcripts and mmRNA molecules.

Abstract: The nucleic acid sequences of adeno-associated virus (AAV) serotype 1 are provided, as are vectors and host cells containing these sequences and functional fragments thereof. Also provided are methods of delivering genes via AAV-1 derived vectors.

Abstract: Methods and compositions are provided for modifying one or more target loci in a cell. Such methods comprise providing a cell comprising a first polynucleotide encoding a first selection marker operably linked to a first promoter active in the cell, wherein the first polynucleotide further comprises a first recognition site for a first nuclease agent. A first nuclease agent is introduced into a cell, wherein the first nuclease agent induces a nick or double-strand break at the first recognition site. Further introduced into the cell is a first targeting vector comprising a first insert polynucleotide flanked by a first and a second homology arm that correspond to a first and a second target site located in sufficient proximity to the first recognition site. At least one cell is then identified comprising in its genome the first insert polynucleotide integrated at the target locus.

Abstract: Methods and compositions are provided for generating targeted genetic modifications on the Y chromosome or a challenging target locus. Compositions include an in vitro culture comprising an XY pluripotent and/or totipotent animal cell (i.e., XY ES cells or XY iPS cells) having a modification that decreases the level and/or activity of an Sry protein; and, culturing these cells in a medium that promotes development of XY F0 fertile females. Such compositions find use in various methods for making a fertile female XY non-human mammal in an F0 generation.

Abstract: The present invention relates to a method of producing a non-human, mammalian oocyte carrying a modified target sequence in its genome, the method comprising the steps of introducing into a non-human, mammalian oocyte: (a) a clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated protein 9 (Cas9 protein) or a nucleic acid molecule encoding said Cas9 protein; and (b-i) a target sequence specific CRISPR RNA (crRNA) and a trans-activating crRNA (tracr RNA) or a nucleic acid molecule encoding said RNAs; or (b-ii) a chimaeric RNA sequence comprising a target sequence specific crRNA and tracrRNA or a nucleic acid molecule encoding said RNA; wherein the Cas9 protein introduced in (a) and the RNA sequence(s) introduced in (b-i) or (b-ii) form a protein/RNA complex that specifically binds to the target sequence and introduces a single or double strand break within the target sequence.

Abstract: Described is a method for the enzymatic production of isoprenol using mevalonate as a substrate and enzymatically converting it by a decarboxylation step into isoprenol as well as the use of an enzyme which is capable of catalyzing the decarboxylation of mevalonate for the production of isoprenol from mevalonate. Furthermore described is the use of mevalonate as a starting material for the production of isoprenol in an enzymatically catalysed reaction.

Abstract: The invention relates to a recombinant carboxydotrophic acetogenic microorganism capable of producing one or more products by fermentation of a substrate comprising CO, wherein the microorganism has an increased tolerance to ethanol versus a parental carboxydotrophic acetogenic microorganism. The invention also provides, inter alia, methods for the production of ethanol and one or more other products from a substrate comprising CO using the recombinant carboxydotrophic acetogenic microorganism.

Abstract: The present invention relates to a recombinant microorganism with improved butanol production ability which has an acetyl-CoA synthesis pathway and a butyryl-CoA synthesis pathway, wherein a pathway converting acetyl-CoA to acetate is inhibited and a pathway converting acetyl-CoA to butyryl-CoA is promoted. In addition, the present invention relates to a method for producing butanol using the recombinant microorganism.

Abstract: Provided herein are metabolically-modified microorganisms useful for producing biofuels. More specifically, provided herein are methods of producing high alcohols including isobutanol, 1-butanol, 1-propanol, 2-methyl-1-butanol, 3-methyl-1-butanol and 2-phenylethanol from a suitable substrate.

Abstract: A method of manufacturing 1,4-butanediol through acetyl-CoA, acetoacetyl-CoA, 3-hydroxybutyryl-CoA, crotonyl-CoA, and 4-hydroxybutyryl-CoA by using a microbe and/or a culture thereof, wherein the microbe in the manufacturing method for 1,4-butanediol includes any one of genes among (a) a gene that has a base sequence of sequence number 1, (b) a gene that has a base sequence such that one or more bases are deleted, substituted, or added in a base sequence of sequence number 1, wherein the gene has a base sequence with an identity greater than or equal to 90% with respect to the base sequence of sequence number 1, and (c) a gene that hybridizes with a gene that has a base sequence complementary with a gene that has a base sequence described in sequence number 1 on a stringent condition, and includes any one or more genes among (d) genes that have base sequences of sequence numbers 2 to 9, (e) genes that have base sequences such that one or more bases are deleted, substituted, or added in base sequences of seque

Abstract: Methods of making C4+ hydrocarbon feedstocks using anaerobic microbes are described.

Abstract: Enzymes for producing non-straight-chain fatty acids, microorganisms comprising the enzymes, and in vivo and in vitro uses of the enzymes. Provided are enzymes capable of producing various non-straight-chain fatty acids, including branched-chain fatty acids, cyclic fatty acids, and furan-containing fatty acids. The enzymes include RSP2144, RSP1091, and RSP1090 from Rhodobacter sphaeroides and homologs thereof. The enzymes can be purified to produce non-straight-chain fatty acids in vitro or expressed in microorganisms to produce non-straight-chain fatty acids in vivo. The microorganisms can be fine-tuned to produce a specific type of non-straight-chain fatty acid by expressing, overexpressing, or deleting the enzymes in various combinations.

Abstract: The present invention is related to biosynthetic oil compositions and methods of making thereof. In some embodiments, the invention relates to the use of endogenous enzymes in plants capable of synthesizing oil. In preferred embodiments, said plants are algae. In further embodiments, said algae are from the family Chlamydomonas, Nannochloropsis, Dunaliella, Chiarella and Scenedesmus. In still further embodiments, said endogenous enzymes are diacylglycerol acyltransferases.

Abstract: A method for constructing an ALA production bacterial strain, the method enhances the activity of related enzymes promoting the synthesis of oxaloacetate and in the 5-aminolevulinic acid (ALA) production bacterial strain, or introducing exogenous related enzymes promoting the synthesis of oxaloacetate, such as phosphoenolpyruvate carboxylase or pyruvate carboxylase, and/or reducing the activity of related enzymes in the downstream metabolic pathway of succinyl coenzyme A in the bacterial strain, such as succinyl coenzyme A synthetase or succinate dehydrogenase, and/or reducing the activity of phosphoenolpyruvate carboxylated kinase and/or malic enzyme. An ALA high-yield bacterial strain constructed by utilizing the method, and method for utilizing the bacterial strain to prepare ALA.

Abstract: A variety of betaine esters, including dialkylaminoalkyl cocoate betaines and dialkylaminoalkyl hydrogenated cocoate betaines are disclosed. These betaines can be advantageously prepared in high yield and purity by a three-step transiterification chemoenzymatic process or a two-step direct esterficiation chemoenzymatic process. These betaine esters have excellent surfactant properties.

Abstract: The present invention relates to a recombinant nucleic acid molecule, a recombinant microorganism, to a method for producing alanine and to the use of the recombinant nucleic acid molecule or the recombinant microorganism for the fermentative production of alanine.

Abstract: A spherical bioreactor system and a method of synthesizing taxol from hazel (Corylus avellana) cell culture are disclosed. The bioreactor has a round bottomed flask fitted with two piezoelectric transducers connected to an audio amplifier, a voltage controlled oscillator, an integrator and a lock in amplifier. For synthesizing taxol, A rapid growing cell line is established from the hazel seeds collected from a cell line culture in a modified MS medium. A suspension culture is established from the rapid growing cell line. The preliminary studies are conducted for estimating an ultrasound dosage and exposure time for treating the suspension culture. The cells are harvested, washed and transferred to a fresh culture media for 1 week. The cells and the culture media are analyzed for molecular and biochemical parameters. The cells are dried for extracting intracellular and extracellular taxols.

Abstract: The present invention provides a process for the preparation of enantiomerically enriched, deuterated secondary alcohols of Formula 1-A by employing ketoreductases or carbonyl reductases without reducing deuterium incorporation.

Abstract: [Problem] To provide a bacterial cellulose which is highly dispersible in a liquid, shows excellent molding properties and high miscibility with other materials when applied to materials, and, therefore, has a high applicability as a practical material, and a bacterium which produces the bacterial cellulose. [Solution] A bacterial cellulose, water that contains said bacterial cellulose at a final concentration of 0.1±0.006 (w/w) showing a light transmittance at a wavelength of 500 nm of 35% or greater, and a bacterium producing the bacterial cellulose. According to the present invention, the bacterial cellulose that is uniformly dispersible in a liquid such as water can be obtained. The bacterial cellulose shows excellent molding properties and high miscibility with other materials and, therefore, can contribute to the improvement in the qualities of a final product or production efficiency thereof or to the reduction of production cost.

Abstract: The present invention provides improved methods for producing a solution containing high molecular weight isolated Streptococcus pneumoniae capsular polysaccharides having phosphodiester linkages between saccharide repeat units. In certain methods, CO2 is supplied to a fermentation culture of Streptococcus pneumoniae bacterial cells that produce capsular polysaccharide serotypes containing phosphodiester linkages between saccharide repeat units. Exemplary Streptococcus pneumoniae serotypes containing a phosphodiester linkage between saccharide repeat units include serotypes 6A, 6B, 19A, and 19F. Supplying CO2 to the fermentation culture includes adding bicarbonate ions to the fermentation culture, adding carbonate ions to the fermentation culture, adding mixtures of bicarbonate and carbonate ions to the fermentation culture, and overlaying the fermentation culture with CO2.

Abstract: A fungal alpha-amylase is provided from Aspergillus terreus (AtAmy1). AtAmy1 has an optimal pH of 4.5 and is operable at 30 75 degrees C., allowing the enzyme to be used in combination with a glucoamylase and an isoamylase in a saccharification reaction. This obviates the necessity of running a saccharification reaction as a batch process, where the pH and temperature must be readjusted for optimal use of the alpha-amylase or glucoamylase. AtAmy1 also catalyzes the saccharification of starch substrates to an oligosaccharide composition significantly enriched in DP2 and (DP1+DP2) compared to the products of saccharification catalyzed by an alpha amylase from Aspergillus kawachii. This facilitates the utilization of the ligosaccharide composition by a fermenting organism in a simultaneous saccharification and fermentation process, for example.

Abstract: A method of producing sugar solution by repeating a sugar solution production process includes steps (1) to (3), wherein a wash solution obtained in step (4) of washing a separation membrane after step (3) is used for step (1) of subsequent sugar solution production processes: step (1): preparing slurry of pretreated cellulose-containing biomass; step (2): hydrolyzing the slurry of pretreated cellulose-containing biomass in step (1) using cellulase from filamentous fungi; and step (3): subjecting the hydrolyzate of step (2) to solid-liquid separation into a solution component and a hydrolysis residue, filtering the solution component through an ultrafiltration membrane and recovering the cellulase from filamentous fungi as a non-permeate, and recovering the sugar solution as a permeate.

Abstract: This disclosure describes unique single stranded DNA templates having a characteristic sequence and secondary structure. The DNA templates disclosed herein are useful for making small RNA molecules through promoterless transcription by a mammalian RNA polymerase, and can serve as an effective vector for producing small RNA molecules of interest in vitro, in situ and in vivo in mammalian cells.

Abstract: The present invention realtes to systems and methods for amplifying nucleic acid. In particular, systems and methods are provided for inhibiting polymerase based terminal transferase activity within a polynucleotide amplification setting (e.g., polymerase chain reaction). In addition, systems and methods are provided for generating amplified products generated with polynucleotide amplification techniques having reduced 3? non-templated nucleotide addition.

Abstract: Disclosed are methods and kits for controlling the length of overhangs at the ends of double stranded DNA based on a competitive inhibition. Further provided are ligase-independent methods for joining the two ends of DNA strands using the same. The present methods efficiently control the overhangs of ds DNA. This user-controlled overhangs supply a tool for efficient ligation in a ligase independent way and can be advantageously used in DNA recombinant technology such as cloning gene or DNA fragments of interest or PCR products.

Abstract: The present invention relates to a new cell-free translation system. In particular, the invention relates to a cell-free reaction system for translating in vitro a RNA into a protein, said reaction system comprising a ribosome-depleted red blood cell lysate and ribosomes isolated from eukaryotic cells, with the proviso that (1) when the ribosome-depleted red blood cell lysate is obtained from a nuclease untreated rabbit reticulocyte lysate, the eukaryotic cells from which ribosomes are isolated are not nuclease untreated rabbit reticulocytes, and (2) when the ribosome-depleted red blood cell lysate is obtained from a nuclease treated rabbit reticulocyte lysate, the eukaryotic cells from which ribosomes are isolated are not nuclease treated rabbit reticulocytes. The invention also pertains to a method for translating in vitro a ribonucleic acid template into an amino acid sequence of interest using the cell-free reaction system of the invention.

Abstract: Herein is reported a method for producing a polypeptide in a prokaryotic cell, comprising the step of cultivating a prokaryotic cell comprising one or more nucleic acids encoding the polypeptide in a chemically defined minimal growth medium and recovering the polypeptide from the prokaryotic cell or the periplasm of the prokaryotic cell or from the medium, wherein the prokaryotic cell is an amino acid auxotrophy cured prokaryotic cell, wherein growth of the amino acid auxotrophy cured prokaryotic cell compared to the non-cured prokaryotic cell under the same cultivation conditions and in the same growth medium requires the supplementation of fewer amino acids to the growth medium, and wherein the chemically defined minimal growth medium is free of the amino acid corresponding to the auxotrophy that has been cured in the auxotrophy cured prokaryotic cell.

Abstract: In situ optical analysis of bacterial endospores can be inhibited when the endospores are present within an optically active carrier medium. To help isolate the endospores from the carrier medium, in some examples, the carrier medium is passed through a hydrophobic material that captures the endospores via hydrophobic attraction. Subsequently, a germination fluid and lanthanide source, such as terbium, can be added to the bacterial endospores captured on the hydrophobic material to form a lanthanide-dipicolinic acid complex in the germination fluid. The germination fluid can then be optically analyzed by measuring the fluorometric response of the lanthanide-dipicolinic acid complex to determine a concentration of the bacterial endospores in the carrier medium.

Abstract: The present invention describes a method to generate new chemical entities (NCEs) that have well-defined activities such as, but not limited to, anti-bacterial, antifungal and anthelmintic effects. The NCEs are generated through adaptive evolution of one microbe (the producer) against another organism or cell type (the target). The producer is made to compete against the target over time by co-culturing the two together and serially passing the producer organism until the producer adaptively evolves by synthesizing an NCE(s) that inhibits growth of or kills the target. The molecular structure of the chemical entity (or entities) is then elucidated using tools from genomics, molecular biology, computational biology, analytical chemistry, organic chemistry and related fields.

Abstract: The present invention is directed to a method for separating, characterizing and/or identifying microorganisms in a test sample. The method of the invention comprises an optional lysis step for lysing non-microorganism cells that may be present in a test sample, followed by a subsequent separation step. The method may be useful for the separation, characterization and/or identification of microorganisms from complex samples such as blood-containing culture media. The invention further provides for the use of one or more identifier agents and interrogating the microorganism sample and/or said one or more identifier agents to produce measurements which characterizing and/or identifying the microorganism based on the produced measurements and/or the presence or absence of the identifier agent or a metabolized form of the identifier agent in the microorganism sample.

Abstract: The current disclosure provides for specific peptides, and derived ionization characteristics of the peptides, from the ALK, Ros, Ron, Ret, TS, and/or FGFR1 proteins that are particularly advantageous for quantifying the ALK, Ros, Ron, Ret, TS, and/or FGFR1 proteins directly in biological samples that have been fixed in formalin by the methods of Selected Reaction Monitoring (SRM) mass spectrometry, or as Multiple Reaction Monitoring (MRM) mass spectrometry. Such biological samples are chemically preserved and fixed wherein the biological sample is selected from tissues and cells treated with formaldehyde containing agents/fixatives including formalin-fixed tissue/cells, formalin-fixed/paraffin embedded (FFPE) tissue/cells, FFPE tissue blocks and cells from those blocks, and tissue culture cells that have been formalin fixed and or paraffin embedded.

Abstract: The present disclosure provides a method for analyzing glycan-derived monosaccharides in a sample. The present disclosure also provides a method for detecting or monitoring a disease or disorder in a patient. In addition, the present disclosure provides a method of determining aberrant glycotransferase activity. The present disclosure further provides a system for analyzing or comparing glycates in a sample.

Abstract: Disclosed herein are compounds useful for detecting oxidants in a living cell, in a multicellular organism, or in a cell-free sample. In particular, disclosed herein are bioluminescent reporter compounds, and more particularly, fluorinated peroxy-caged-luciferin (PCL) compounds, compositions comprising such compounds, methods of using such compounds and compositions, and processes for preparing such compounds. Also disclosed herein are kits and methods for detecting and measuring peroxynitrite, and optionally, additional oxidants in a living cell, in a multicellular organism, or in a cell-free sample.

Abstract: The present invention provides for novel methods and compositions for nucleic acid sequence detection. Unique, identifying cleavage fragments from probes, bound to target nucleic acids, are produced during PCR by the 5?-nuclease activity of the polymerase. The identity of the targets can be determined by identifying the unique cleavage fragments.

Abstract: Systems and methods for preventing or reducing unwanted heat in a microfluidic device while generating heat in selected regions of the device are described. Current can be supplied to a heating element through electric leads that are designed so that the current density in the leads is substantially lower than the current density in the heating element. Unwanted heat in the microfluidic complex can be reduced by thermally isolating the electric leads from the microfluidic complex by, for example, running each lead directly away from the microfluidic complex. Unwanted heat can be removed from selected regions of the microfluidic complex using one or more cooling devices.

Abstract: Compositions and methods for detecting Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) are provided. The present invention also provides methods and compositions for screening for infection/inflammation based on genomic copy number. Described herein is a method that entails assaying a sample obtained from the urogenital tract of the mammal for an indicator of genomic copy number, wherein a genomic copy number level that is higher than a control genomic copy number level is indicative of the presence of infection or inflammation of the urogenital tract.

Abstract: A test kit for rapidly detecting a live bacterium is disclosed. The test kit for rapidly detecting a live bacterium includes ethidium monoazide, a magnetic bead, a first primer pair and a microfluidic chip. A test kit for rapidly determining an appropriate antibiotic is also disclosed. The test kit for rapidly determining an appropriate antibiotic includes ethidium monoazide, a magnetic bead, four primer pairs and a microfluidic chip.

Abstract: The present disclosure describes a method for identifying a strain or species of bacteria using a single locus sequence typing technique. The single locus useful in the method is the promoter region of the 16S rRNA operon. The method is useful to identify infectious bacteria in a subject, to identifying contaminants in a food source, as well as strain typing and genetic fingerprinting of bacterial families.

Abstract: This invention relates to a rapid method for detection and characterization of Escherichia coli bacteria serotype O157:H7 based on the presence of nucleic acid sequences, in particular, to a PCR-based method for detection, and to oligonucleotide molecules and reagents and kits useful therefore. This method is preferably employed to detect E. coli O157:H7 in a food or water sample, such as a beef enrichment. The present invention further relates to replication compositions and kits for carrying out the method of the present invention.

Abstract: Disclosed herein are methods, compositions and kits for the quantification of a nucleic acid target present on a solid support. This entails quantitative real-time polymerase chain reaction wherein minor groove binders are excluded.

Abstract: This invention provides novel compositions and processes for analyte detection, quantification and amplification. Nucleic acid arrays and libraries of analytes are usefully incorporated into such compositions and processes. Universal detection elements, signaling entities and the like are employed to detect and if necessary or desirable, to quantify analytes. Amplification of target analytes are also provided by the compositions and processes of this invention.

Abstract: Provided herein are dynamic flux nucleic acid sequence amplification methods. The dynamic flux nucleic acid sequence amplification methods described herein are capable of amplifying nucleic acid sequences within a narrow temperature range. In some aspects, the disclosure provides for real-time dynamic flux nucleic acid sequence amplification methods.

Abstract: Methods, systems and compositions where a target nucleic acid includes a registration sequence disposed therein for identification of the number or relative position of determined sequence from the template sequence. Particularly preferred aspects include a registration sequence in a circular template nucleic acid sequence which is, in turn, used in sequence by incorporation processes that rely upon template dependent, polymerase mediated primer extension in the identification of the sequence of the template.

Abstract: Massively parallel sequencing of cell-free, maternal plasma DNA was recently demonstrated to be a safe and effective screening method for fetal chromosomal aneuploidies. Here, we report an improved sequencing method achieving significantly increased throughput and decreased cost by replacing laborious sequencing library preparation steps with PCR employing a single primer pair. Using this approach, samples containing as little as 4% trisomy 21 DNA could be readily distinguished from euploid samples.

Abstract: Provided herein are devices and methods suitable for sequencing, amplifying, analyzing, and performing sample preparation procedures for nucleic acids and other biomolecules.

Abstract: A method of selecting nucleic acid samples including particular desired alleles from a plurality of nucleic acid samples including the steps of performing a first reaction in a plurality of pools containing the samples to produce reaction products including a source tag identifying said each pool; pooling the pools to provide pooled pools; for each of the desired alleles to be identified, performing a second reaction using said reaction products to produce allele-specific second reaction products comprising a marker tag and a derived source tag; identifying said allele-specific second reaction products, and further identifying nucleic acid samples with the desired alleles. A source tag sharing number “d” may be determined for each of the alleles. Alleles may also be binned together.

Abstract: An analytical assembly within a unified device structure for integration into an analytical system. The analytical assembly is scalable and includes a plurality of analytical devices, each of which includes a reaction cell, an optical sensor, and at least one optical element positioned in optical communication with both the reaction cell and the sensor and which delivers optical signals from the cell to the sensor. Additional elements are optionally integrated into the analytical assembly. Methods for forming and operating the analytical system are also disclosed.