Abstract: Provided herein are constructs for genome editing or genetic engineering its fungi or protists, methods of using the constructs and media for use in selecting cells. The construct include a polynucleotide encoding a thymidine kinase operably connected to a promoter, suitably a constitutive promoter; a polynucleotide encoding an endonuclease operably connected to an inducible promoter; and a recognition site for the endonuclease. The constructs may also include selectable markers for use in selecting recombinations.

Abstract: The present invention is directed to antibodies and fragments thereof and humanized versions thereof having binding specificity for IL-6. Another embodiment of this invention relates to the antibodies described herein, and binding fragments thereof, comprising the sequences of the VH, VL and CDR polypeptides described herein, and the polynucleotides encoding them. The invention also contemplates conjugates of anti-IL-6 antibodies and binding fragments thereof conjugated to one or more functional or detectable moieties. The invention also contemplates methods of making said anti-IL-6 antibodies and binding fragments thereof. Embodiments of the invention also pertain to the use of anti-IL-6 antibodies, and binding fragments thereof, for the diagnosis, assessment and treatment of diseases and disorders associated with IL-6.

Abstract: Compositions, peptide solutions and macroscopic scaffolds of self-assembling peptides consisting essentially of non-ionic, polar amino acids are provided. Particular peptides include those comprising or consisting essentially of serine, threonine, tyrosine, cysteine, glutamine, asparagine, methionine, tryptophan, hydroxy-proline, and combinations thereof. Methods of sterilizing the self-assembling peptides, and scaffolds comprising the peptides are also provided.

Abstract: This invention relates generally to the discovery of novel recombinant forms of ?-hexosyl-transferases (BHT) and uses thereof to produce galacto-oligosaccharides (GOS) or as food additives.

Abstract: Methods for producing heterologous multi-subunit proteins in transformed cells are disclosed. In particular, the present disclosure provides improved methods of producing multi-subunit proteins, including antibodies and other multi-subunit proteins, which may or may not be secreted, with a higher yield and decreased production of undesired side-products. In exemplary embodiments, the transformed cells are a yeast, e.g., methylotrophic yeast such as Pichia pastoris.

Abstract: Provided is an adjuvant composition comprising a nucleic acid comprising a double-stranded RNA bound to a single-stranded DNA, the double-stranded RNA consisting of a nucleotide sequence of SEQ ID NO: 1 and its complementary sequence, the single-stranded DNA consisting of a nucleotide sequence of SEQ ID NO: 2. Also provided is a vaccine composition comprising the adjuvant composition and an antigen or an antigen component. The nucleic acid contained in the adjuvant composition is a chemically synthesizable nucleic acid having potent adjuvant activity and high safety.

Abstract: The present invention is directed to compositions methods and kits for regulation of gene therapies, including, without limitation, reversible gene therapies and allele-specific therapies.

Abstract: Described herein are methods and genetically engineered cells useful for producing an altered N-glycosylation form of a target molecule. Also described are methods and molecules with altered N-glycosylation useful for treating a variety of disorders such as metabolic disorders.

Abstract: The present invention relates to a method for mass-producing an antifreeze protein derived from a polar yeast, and more particularly, to a method for mass-producing an antifreeze protein derived from Leucosporidium sp., which is the polar yeast, for synthesizing a recombinant polynucleotide by optimizing and altering a gene, which codes the antifreeze protein derived from the polar yeast, for a yeast expression system, and for expressing same using the yeast expression system.

Abstract: The present invention relates to a methodology for the generation of infectious ribonucleoparticles (RNPs) of negative-strand RNA viruses, and in particular of non-segmented negative-strand RNA viruses in yeast, especially in budding yeast. Accordingly, the patent application relates to a recombinant yeast strain suitable for the rescue of infectious non-segmented negative-strand RNA virus particles or infectious virus-like particles. The invention also relates to the use of the recombinant yeast to prepare vaccine seed and to the use of the produced RNPs or RNPs-like to prepare vaccine formulations. It also concerns the use of the recombinant yeast for the screening of libraries of DNA.

Abstract: The invention refers to a method of producing a recombinant polypeptide of interest (POI) in a cell culture, comprising genetically engineering a eukaryotic cell line—to specifically cause prolongation of the G2+M cell cycle phase in a pre-culture phase, and—to produce the POI in a producing phase following the pre-culture phase, a high producer cell line and cell culture as well as a method of increasing the yield of a recombinant POI production in a cell culture.

Abstract: This invention relates to protein and/or polypeptide production, particularly improved production of extracellular heterologous polypeptides and proteins. In particular, the invention relates to compositions of cell populations capable of improved levels of extracellular secretion relative to control populations, kits containing such compositions, methods of producing heterologous proteins of interest and recombinant microorganisms capable of improved extracellular heterologous protein production.

Abstract: The present invention relates to a method of expressing an immunotoxin in Pichia pastoris strain mutated to toxin resistance comprising a) growing the Pichia pastoris in a growth medium comprising an enzymatic digest of protein and yeast extract and maintaining a dissolved oxygen concentration at 40% and above; and b) performing methanol induction with a limited methanol feed of 0.5-0.75 ml/min/IO L of initial volume during induction along with a continuous infusion of yeast extract at a temperature below 17.5° C., antifoaming agent supplied up to 0.07%, agitation reduced to 400 RPM, and the induction phase extended out to 163 h.

Abstract: In accordance with the present invention, a method for increasing the yield of rLV vector particles comprising a trans gene encoding a therapeutic protein or fragment thereof is disclosed. In one approach, cells are transfected with plasmids encoding the necessary components for rLV production using a calcium chloride transfection mix at pH 7.1 wherein the calcium chloride and plasmids form a complex which is added to the cells at a constant speed. The cells are then incubated for a suitable time period wherein virus particle media is collected at least twice during the incubation period and stored in a cold storage unit, thereby reducing virus inactivation.

Abstract: A selectively inducible, single-stranded DNA (ssDNA) expression library, a method for constructing a ssDNA expression library, a method for screening ssDNA using the expression library, and a method for identifying ssDNA molecules that alter expression of bacterial and fungal gene(s) related to cell growth and toxin production and secretion. The screening library is used to, among other things, identify ODNs effective in stopping cell growth, killing bacteria or fungi, or preventing bacteria and/or fungi from synthesizing and secreting their toxins, and/or to discover ODNs effective in eukaryotic (e.g., mammalian) cells for targeted alteration of gene function. The library is also useful for identifying ssDNAs or ODNs that are used as therapeutic agents for, for instance, providing a method for treatment of bacterial infections such as sepsis.

Abstract: The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities such as those involved in glycosylation to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified oligosaccharides are created or selected. N-glycans made in the engineered host cells have a Man5GlcNAc2 core structure which may then be modified further by heterologous expression of one or more enzymes, e.g.

Abstract: The present invention is directed to a method for preparing an expression vector encoding a tailored recombinase, wherein said tailored recombinase recombines asymmetric target sites within the LTR of proviral DNA of a retrovirus inserted into the genome of a host cell and is useful as means for excising the provirus from the genome of the host cell. The present invention further relates to an in vitro-method of optimising the treatment of a retroviral infection of a subject and to the use of tailored recombinases for the preparation of pharmaceutical compositions for reducing the viral load in a subjected infected by a retrovirus.

Abstract: The present invention relates to non-catalytic carbohydrate-binding modules (CBM) belonging to a new family of CBM's. A CBM of the invention was found attached to a glycosyl hydrolase family 61 (GH61) polypeptide and was shown to have little homology with known CBM's indicating that it is the first known member of a new family of CBM's. The present invention further relates to CBM's preferably exhibiting binding affinity for cellulose; to a method of producing such CBM's; and to methods for using such CBM's in the textile, detergent and cellulose fiber processing industries, for purification of polypeptides, immobilization of active enzymes, baking, manufacturing of biofuel, modification of plant cell walls.

Abstract: The present invention relates to a gene associated with ethanol tolerance, and yeast strains and uses using the same. The yeast strain of this invention may growth under the condition not only with high-concentration ethanol, preferably 6-15% ethanol, but also in high osmotic pressure, preferably 30-40% glucose or sucrose. The present inventors developed yeast strains resistant to high-concentration glucose and ethanol, suggesting that they would be valuably applied to much effective ethanol production, and also be utilized as a superbacteria having tolerance to various stresses for ethanol production with high efficiency.

Abstract: The invention relates to methods of separation and/or purification of impurities yielding a purified heterologous protein product devoid of related impurities or with substantially minimal quantities of such glycosylated impurities. More specifically, the invention relates to the identification of glycosylated forms of insulin analogues such as glargine impurities characterized post expression in yeast based systems such as Pichia pastoris. The invention also relates to methods used to clone gene encoding the protein insulin glargine; inserting the related gene in a suitable yeast host; producing culture of the recombinant strain, stimulating expression of the heterologous polypeptide, its secretion and purification post fermentation and related enzymatic conversions.

Abstract: The subject of the present invention is a process for preparing a genetically modified yeast by multicopy integration of at least four expression cassettes, allowing the production of a molecule of interest at high titre. The subject of the present invention is also yeasts transformed according to said process, and the use thereof for producing hydrocortisone.

Abstract: The technology relates in part to biological methods for producing adipic acid and engineered microorganisms capable of such production.

Abstract: Sugars comprising the monosaccharides glucose and fructose, and the disaccharides sucrose and mannose are catalytically converted to ethanol in a sulfate fortified acid medium in the presence of transition metal compounds possessing a degree of symmetry. This is not a fermentation process but is a catalytic chemical process where conversion efficiency is improved by saturating the acidic reaction mixture with inorganic sulfate salts to reduce competitive reactions. Ethanol formed during the reaction is removed by distillation facilitating a continuous process.

Abstract: The present invention provides methods of modifying in vivo mutagenesis or homologous and homeologous recombination in a eukaryote. The method of modifying in vivo mutagenesis involves transforming a eukaryote with a nucleotide sequence capable of expressing a wild-type prokaryotic MutS, MutL, MutH, MutU, NLS-MutS, NLS-MutL, NLS-MutH, NLS-MutU protein, or a combination thereof, and expressing the protein. A method of modifying recombination between homologous chromosomes in an allopolyploid eukaryotic organism comprising, expressing a nucleotide sequence encoding prokaryotic NLS-MutS in combination with one or more than one of NLS-MutL, NLS-MutH, or NLS-MutU, within a germ cell of the allopolyploid eukaryotic organism is also disclosed.

Abstract: The invention provides a yeast strain and a method for making the same. The method has the step of replacing the regulation region upstream of the hsp104 gene in the genome of the yeast, so as to accelerate and prolong the expression span of hsp104 gene and enhance the capability of the yeast to ferment and produce ethanol in a high-temperature environment. The yeast is capable of fermenting glucose at a temperature higher than 42° C. to produce ethanol, or biomass ethanol, wherein the ethanol production ratio based on fermentation of glucose is higher than 97%. Being able to synchronize the degradation/hydrolysis stage and fermentation stage of biomass ethanol producing process, the yeast in accordance with the present invention is able to lower the production cost of biomass ethanol and further raise the productivity with its high ethanol production ratio.

Abstract: The present application relates to genetically modified yeasts for the production of glycoproteins having optimized and homogeneous glycan structures. These yeasts comprise an inactivation of the Och 1 gene, the integration by homologous recombination, into an auxotrophic marker, of an expression cassette comprising a first promoter, and an open reading frame comprising the coding sequence for an ?-1,2-mannosidase I, and the integration of a cassette comprising a second promoter different from said first promoter and the coding sequence for an exogenous glycoprotein. These yeasts make it possible to produce EPO with an optimized and 98% homogeneous glycosylation.

Abstract: The invention relates to a strain of the yeast Saccharomyces cerevisiae which, owing to deletion of the genomic sequences, no longer synthesizes hexose transporters and, as a consequence, can no longer grow on substrates with hexoses as the only carbon source, and whose ability of growing on a substrate with a hexose as the only carbon source is restored when it expresses a GLUT4 gene.

Abstract: The present invention provides a method for producing an yeast having an increased cellulose hydrolysis ability. The method includes the step of introducing increased integration copy numbers of both a gene for an enzyme capable of hydrolyzing crystalline cellulose and a gene for an enzyme capable of hydrolyzing noncrystalline cellulose into a noncellulolytic yeast to give a transformed yeast. The yeast having an increased cellulose hydrolysis ability can be suitably used for ethanol production from cellulose-based materials.

Abstract: Coronatine has been found to enhance binding of the JAZ1 degron to the Arabidopsis F-box protein COI1, and analysis of the JAZ1 degron sequence has resulted in the identification of specific peptide sequences that bind COI1 with high affinity in the presence of coronatine. Crystal structure analysis has determined that coronatine and JA-Ile enhance the interaction between COI1 and JAZ1 via binding to a specific binding pocket on COI1. Attachment of one or more JAZ1 peptide tags as disclosed herein to a target protein in a non-plant cell expressing Arabidopsis COI1 or a homolog thereof results in degradation of the target protein following addition of a molecule that binds the coronatine/JA-Ile binding pocket on COI1. Therefore, provided herein are compositions, methods, and kits for targeted protein degradation.

Abstract: Methods and compositions are described that relate to obtaining concentrated preparations of secreted recombinant proteins. These proteins are expressed in the form of fusion proteins with a chitin-binding domain (CBD). The fusion proteins are capable of being concentrated in the presence of chitin. Also described is: a shuttle vector that includes a modified LAC4 promoter; a chitinase-negative host cell; a CBD capable of eluting from chitin under non-denaturing conditions; and sterilized chitin, which can be optionally magnetized for facilitating recovery of recombinant protein.

Abstract: The present invention includes a method to produce a recombinant mite Group 1 protein in a methyltrophic yeast or an Escherichia coli microorganism. The present invention also relates to a recombinant mite Group 1 protein obtained by such a method, such a recombinant protein being able to selectively bind IgE or cause proliferation of a T cell that proliferate in response to a native mite Group 1 protein. Also included in the present invention is the use of such a recombinant mite Group 1 protein to detect mite allergy or to reduce an allergic response to a mite Group 1 protein. The present invention also includes novel mite Group 1 nucleic acid molecules, proteins, recombinant molecules, and recombinant cells, as well as uses thereof.

Abstract: The invention provides molecular switches which couple external signals to functionality, and combinatorial methods of making and using the same involving circular permutation of nucleic acid and amino acid sequences. The switches according to the invention can be used, for example, to regulate gene transcription, target drug delivery to specific cells, transport drugs intracellularly, control drug release, provide conditionally active proteins, perform metabolic engineering, and modulate cell signaling pathways. Libraries comprising the switches, expression vectors and host cells for expressing the switches are also provided.

Abstract: Increasing tolerance to butanol in yeast has been accomplished by increasing activity of the high osmolarity/glycerol response pathway. Yeast with increased expression of PBS2p, a mitogen activated protein kinase kinase of the MAPK module of the high osmolarity/glycerol response pathway may be used for improved butanol production.

Abstract: A method is presented for selecting and isolating nucleic acids capable of conferring tolerance or resistance to environmental stress conditions in plants or yeast. Furthermore, nucleic acids, the proteins they encode and their use for the production of plants or yeast with enhanced environmental stress resistance is disclosed.

Abstract: The present invention provides genetically engineered strains of Pichiacapable of producing proteins with reduced glycosylation. In particular, the genetically engineered strains of the present invention are capable of expressing either or both of an ?-1,2-mannosidase and glucosidase II. The genetically engineered strains of the present invention can be further modified such that the OCH1 gene is disrupted. Methods of producing glycoproteins with reduced glycosylation using such genetically engineered stains of Pichia are also provided.

Abstract: The present invention relates to a new group of pesticides consisting of inactivated microorganisms containing double-strand RNA molecules (dsRNA), corresponding to receptor genes coupled with G proteins (GPCR) whose functioning is vital for phytophagous invertebrates (insects, mites and molluscs) or for infesting or in any case harmful organisms for the health of human beings and domestic animals, and to a method for the preparation of said microorganisms.

Abstract: The technology relates in part to biological methods for producing adipic acid and engineered microorganisms capable of such production.

Abstract: The present invention relates to a cell comprising an Endoplasmic Reticulum (ER) stress element operably linked to a reporter element and an exogenous gene encoding a protein that induces ER stress. Methods of screening using the modified cell, constructs used in the modified cell, the candidate agents identified by the screen and uses thereof are also part of the invention.

Abstract: The present invention provides compositions and methods for rapid assembly of one or more assembled polynucleotides from a plurality of component polynucleotides. The methods of the invention utilize circular nucleic acid vectors that comprise a DNA segment D flanked by an annealable linker sequence, annealable linker sequence pairs LA and LB, or annealable linker sequence/primer binding segment pairs LA and PB or PA and LB. Restriction endonuclease digestion of a plurality of vectors containing the DNA segments to be assembled generates a plurality of DNA fragments comprising the elements PA-D-LB, LA-D-LB, and LA-D-PB or D-LB, LA-D-LB, and LA-D. The sequences of annealable linker sequences LA and LB provide complementary termini to the DNA fragments, which are utilized in host cell mediated homologous recombination or together with promer binding segments PA and PB in a polymerase cycling assembly reaction for the ordered assembly of the various DNA segments into one or more assembled polynucleotides.

Abstract: Sucrose biosensors are disclosed, which comprise a sucrose binding domain conjugated to donor and fluorescent moieties that permit detection and measurement of Fluorescence Resonance Energy Transfer upon sucrose binding. Such biosensors are useful for real time monitoring of sucrose metabolism in living cells.

Abstract: Disclosed are the ADE3, ADE4, ADE5, 7, ADE6, ADE8, ADE12, and ADE13 genes encoding various enzymes in the adenine biosynthesis pathway of Pichia pastoris. The loci in the Pichia pastoris genome encoding these enzymes are useful sites for stable integration of heterologous nucleic acid molecules into the Pichia pastoris genome. The genes or gene fragments encoding the particular enzymes, which may be used as selection markers for constructing recombinant Pichia pastoris.

Abstract: Disclosed are the LYS1, LYS2, LYS4, LYS5, and LYS9 genes encoding various enzymes in the lysine biosynthesis pathway of Pichia pastoris. The loci in the Pichia pastoris genome encoding these enzymes are useful sites for stable integration of heterologous nucleic acid molecules into the Pichia pastoris genome. The genes or gene fragments encoding the particular enzymes may be used as selection markers for constructing recombinant Pichia pastoris.

Abstract: Disclosed are the URA1, URA2, URA4, and URA6 genes encoding various enzymes in the uracil biosynthesis pathway of Pichia pastoris. The loci in the Pichia pastoris genome encoding these enzymes are useful sites for stable integration of heterologous nucleic acid molecules into the Pichia pastoris genome. The genes or gene fragments encoding the particular enzymes may be used as selection markers for constructing recombinant Pichia pastoris.

Abstract: Disclosed is the HIS7 gene encoding the His7p enzyme in the histidine biosynthesis pathway of Pichia pastoris. The locus in the Pichia pastoris genome encoding the His7p is useful sites for stable integration of heterologous nucleic acid molecules into the Pichia pastoris genome. The gene or gene fragment encoding the His7p may be useful as a selection marker for constructing recombinant Pichia pastoris.

Abstract: Disclosed are the MET1, MET3, MET4, MET6, MET7, MET8, MET10, MET14, MET16, MET17, MET19, MET22, MET2, and MET28 genes encoding various enzymes in the methionine biosynthesis pathway of Pichia pastoris. The loci in the Pichia pastoris genome encoding these enzymes are useful sites for stable integration of heterologous nucleic acid molecules into the Pichia pastoris genome. The genes or gene fragments encoding the particular enzymes may be used as selection markers for constructing recombinant Pichia pastoris.

Abstract: Disclosed are the ARG5, 6, ARG8, ARG9, ARG80, ARG81, and ARG82 genes encoding various enzymes in the arginine biosynthesis pathway of Pichia pastoris. The loci in the Pichia pastoris genome encoding these enzymes are useful sites for stable integration of heterologous nucleic acid molecules into the Pichia pastoris genome. The genes or gene fragments encoding the particular enzymes may be used as selection markers for constructing recombinant Pichia pastoris.

Abstract: A method for producing a chimaeric human papillomavirus (HPV) L1 polypeptide containing a heterologous peptide, and in particular, a HPV L2 peptide comprising the steps of introducing a DNA sequence coding for the heterologous peptide into a DNA sequence coding for the L1 polypeptide; introducing the DNA sequence including the sequences for the L1 polypeptide and heterologous peptide into a host cell in which the DNA sequence can be expressed; causing expression of the DNA sequence; and recovering the resulting chimaeric L1 polypeptide which includes the heterologous peptide. The invention also describes a vector for use in the method, a host cell containing the vector, and a vaccine including the chimaeric HPV L1 polypeptide produced according to the method.

Abstract: The present invention provides methods to reduce or eliminate ?-mannosidase resistant glycans on glycoproteins in yeast. The reduction or elimination of ?-mannosidase resistant glycans on glycoproteins results from the disruption of the newly isolated P. pastoris AMR2 gene encoding ?1,2-mannosyltransferase. The present invention also discloses novel genes, polypeptides, antibodies, vectors and host cells relating to ?-mannosidase resistance on glycans.

Abstract: The present invention relates to recombinant microorganisms comprising biosynthetic pathways and methods of using said recombinant microorganisms to produce various beneficial metabolites. In various aspects of the invention, the recombinant microorganisms may further comprise one or more modifications resulting in the reduction or elimination of 3 keto-acid (e.g., acetolactate and 2-aceto-2-hydroxybutyrate) and/or aldehyde-derived by-products. In various embodiments described herein, the recombinant microorganisms may be microorganisms of the Saccharomyces clade, Crabtree-negative yeast microorganisms, Crabtree-positive yeast microorganisms, post-WGD (whole genome duplication) yeast microorganisms, pre-WGD (whole genome duplication) yeast microorganisms, and non-fermenting yeast microorganisms.

Abstract: This invention relates generally to the discovery of novel recombinant forms of ?-hexosyl-transferases (BHT) and uses thereof to produce galacto-oligosaccharides (GOS) or as food additives.