Abstract: The present invention provides novel methods of producing diphtheria toxin. In particular, the present invention provides novel methods of producing nontoxic forms of diphtheria toxin, e.g., CRM197. The present invention also provides novel compositions comprising diphtheria toxin or nontoxic forms of diphtheria toxin, e.g., CRM197.

Abstract: The present invention relates to a nucleic acid molecule that comprises at least one synthetic GAL1 core promoter containing one or more ligand responsive operator sequences, preferably one or more FadR operator sites; at least one upstream enhancer element (UEE); and at least one nucleotide sequence encoding for one or more genes of interest; wherein the at least one synthetic GAL1 core promoter is operably linked to the at least one nucleotide sequence encoding for one or more genes of interest and the at least one UEE is operably linked to the at least one synthetic GAL1 core promoter. Also encompassed are expression systems and recombinant cells that include these nucleic acid molecules and methods that use these nucleic acid molecules, expression systems or cells.

Abstract: The present disclosure provides compositions and methods for using recombinant C1 metabolizing microorganisms capable of metabolizing sulfur containing compounds and other contaminants to biologically convert sour or acidic natural gas into high-value molecules, and to allow recovery of stranded oil.

Abstract: The present invention provides oligonucleotide constructs, sets of such oligonucleotide constructs, and methods of using such oligonucleotide constructs to provide validated sequences or sets of validated sequences corresponding to desired ROIs. Such validated ROIs and constructs containing these have a wide variety of uses, including in synthetic biology, quantitative nucleic acid analysis, polymorphism and/or mutation screening, and the like.

Abstract: The present invention provides oligonucleotide constructs, sets of such oligonucleotide constructs, and methods of using such oligonucleotide constructs to provide validated sequences or sets of validated sequences corresponding to desired ROIs. Such validated ROIs and constructs containing these have a wide variety of uses, including in synthetic biology, quantitative nucleic acid analysis, polymorphism and/or mutation screening, and the like.

Abstract: The present invention relates to methods of joining two or more double-stranded (ds) or single-stranded (ss) DNA molecules of interest in vitro, wherein the distal region of the first DNA molecule and the proximal region of the second DNA molecule of each pair share a region of sequence identity. The method allows the joining of a large number of DNA fragments, in a predetermined order and orientation, without the use of restriction enzymes. It can be used, e.g., to join synthetically produced sub-fragments of a gene or genome of interest. Kits for performing the method are also disclosed. The methods of joining DNA molecules may be used to generate combinatorial libraries useful to generate, for example, optimal protein expression through codon optimization, gene optimization, and pathway optimization.

Abstract: Methods are provided for the synthesis and secretion of recombinant hetero-multimeric proteins in mating competent yeast. A first expression vector is transformed into a first haploid cell; and a second expression vector is transformed into a second haploid cell. The transformed haploid cells, each individually synthesizing a non-identical polypeptide, are identified and then genetically crossed or fused. The resulting diploid strains are utilized to produce and secrete fully assembled and biologically functional hetero-multimeric protein.

Abstract: A gene expression cassette capable of producing psicose at high yield with high stability, a GRAS (Generally recognized as safe) microorganism, a method of producing the enzyme by using the GRAS microorganism, and a method of producing the psicose by using the GRAS microorganism and enzyme are provided.

Abstract: Disclosed are yeast cells expressing a polypeptide comprising a signal sequence and a human amyloid beta protein. Also disclosed are methods of screening yeast cells to identify compounds that prevent or suppress amyloid beta-induced toxicity and genetic suppressors or enhancers of amyloid beta-induced toxicity. Compounds identified by such screens can be used to treat or prevent neurodegenerative disorders such as Alzheimer's disease.

Abstract: Provided is a ramp tag capable of solving instability in translation rate resulting from poor compatibility between codons in a foreign gene and a host when expressing a recombinant protein in E. coli. Unlike the conventional codon optimization or codon deoptimization method for solving the problem of rare codons, the present invention increases an expression efficiency of a target protein by merely having the ramp tag be fused with a target gene or independently expressed, without changing the original codon sequence, thereby allowing tRNA to be reused. Thus, the present invention provides a novel method for increasing recombinant protein expression which is capable of reducing costs and time in comparison to the codon optimization method that artificially synthesizes DNA sequences. Therefore, it is expected that the method of the present invention will be able to be used in production of high value-added pharmaceuticals or industrial enzymes.

Abstract: Methods for producing heterologous proteins are disclosed. In particular, the present disclosure provides improved methods of producing desired proteins, including multi-subunit proteins such as antibodies, with a higher yield and improved purity. In exemplary embodiments, the transformed cells are a yeast, e.g., methylotrophic yeast such as Pichia pastoris.

Abstract: A method for producing from host cells a heterologous polypeptide or protein comprising at least one cysteine residue in a reduced state. The method cultured the host cells in a culture medium comprising a reducing agent; and recovered the heterologous polypeptide or protein comprising at least one cysteine residue in a reduced state from the host cells or from the culture medium, where the host cells comprise a nucleic acid encoding the heterologous protein or polypeptide, and where the reducing agent is capable of permeating the plasma membrane of the host cells.

Abstract: The invention relates to methods for predicting whether a sample contains enterohemorrhagic Escherichia coli (EHEC) of at least one of EHEC O157:[H7], O145:[H28], O103:[H2], O111:[H8], O121:[H19], O26: [H11], O45:[H2] or O104: [H4] serotypes, and for identifying said serotypes, through detection of gene espK in association with at least one of the genetic markers Z1151, Z1153, Z1154, Z1155, Z1156, Z6065, Z2098, ureD or espV and/or through detection of serotype-specific CRISPR sequences.

Abstract: A feedback control mechanism for a fermentation of yeast cells to make recombinant proteins uses a respiratory quotient measurement which adjusts the levels of oxygenation and/or fermentable sugar feed. The feedback control mechanism permits well controlled cultures that produce good amounts of product while avoiding toxic accumulation of ethanol. Additionally, recombinant proteins so produced have excellent qualitative properties, such as excellent homogeneity and proper inter-subunit assembly.

Abstract: Disclosed are genetically engineered organisms, such as yeast and bacteria, that have the ability to metabolize atypical phosphorus or sulfur sources. Fermentation methods using the genetically engineered organisms are also described. The fermentation methods are robust processes for the industrial bioproduction of a variety of compounds, including commodities, fine chemicals, and pharmaceuticals.

Abstract: Methods are provided for the synthesis and secretion of recombinant hetero-multimeric proteins in mating competent yeast. A first expression vector is transformed into a first haploid cell; and a second expression vector is transformed into a second haploid cell. The transformed haploid cells, each individually synthesizing a non-identical polypeptide, are identified and then genetically crossed or fused. The resulting diploid strains are utilized to produce and secrete fully assembled and biologically functional hetero-multimeric protein.

Abstract: A method of assembling multi-fragment DNA molecule, the method including: 1) constructing a vector system including an acceptor vector A1, a first donor vector B1, and a second donor vector B2, and each of the acceptor vector A1, the second donor vector B2, and the second donor vector B2 including a multiple cloning site; 2) introducing a first target sequence Lm to the first donor vector B1, m being an odd number starting from 1, and introducing a second target sequence Ln to the second donor vector B2, n being an even number starting from 2; and 3) repeating 2) to assemble multiple fragments to yield a multi-fragment DNA molecule.

Abstract: The present invention relates to the production and use of covalently closed circular (ccc) recombinant plasmids, and more particularly to vector modifications that improve expression of said DNA molecules in the target organism.

Abstract: Described are methods for production of RNA transcripts using a non-amplified, linearized DNA template in an in vitro transcription reaction. Enzymatic 5? capping and oligo dT purification can also be included in the methods.

Abstract: Methods for producing heterologous proteins are disclosed. In particular, the present disclosure provides improved methods of producing desired proteins, including multi-subunit proteins such as antibodies, with a higher yield and improved purity. In exemplary embodiments, the transformed cells are a yeast, e.g., methylotrophic yeast such as Pichia pastoris.