Abstract: The present invention relates to a modified Bacillus licheniformis host cell, wherein one or more naturally occurring chromosomal chloramphenicol acetyl transferase encoding gene(s), cat, has been inactivated. The inactivation of the chromosomal cat gene(s) allows the use of chloramphenicol as an efficient selective agent in methods for DNA introduction into the modified cell.

Abstract: Disclosed herein is a genetic modification of moderately thermophilic Bacillus species that are facultative anaerobic and homolactic. The method includes introducing DNA cloned in a thermosensitive plasmid system containing a pSH71 replicon or a homologue thereof into cells of a moderately thermophilic Bacillus species that is facultative anaerobic and homolactic; culturing the cells on a selective medium at a permissive temperature to select transformed cells; culturing the transformed cells on a selective medium at a non-permissive temperature to select transformed cells capable of growing on the selective medium at the non-permissive temperature. The method can modify the Bacilli for R-lactic acid production, production of other organic acids than lactic acid, alcohol, enzymes, amino acids, and vitamins. The Bacillus species may be modified by replacing the S-lactate dehydrogenase gene by a DNA construct including a DNA sequence encoding R-lactate dehydrogenase.

Abstract: The present invention relates to methods of obtaining genetic competence in non-competent Bacillus cells for their transformation with exogenous DNA.

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: A bioengineering, in particular to develop novel probiotic preparations based on Bacillus-strain bacteria, which can be used for preventing and treating infectious diseases and disbiosis of a human being, farm animals and birds. The novel strains of B. subtilis 07 (VKPM No. B-8611) and B. licheniformis 09 (VKPM No. B-8610) exhibit a broad spectrum of antagonistic activity, high proteolytic and amylase activity and a distinct ability in terms of a lysozim production. Such strains do not compete with each other but enter into synergistic relations in the form of an increased antagonistic action of the biopreparation. The inventive biopreparation comprises the B. subtilis 07 VKPM No. B-8611 and B. licheniformis 09 VKPM No. B-8610 strains and a protective medium. Such biopreparation can also contain a solvent and/or filler and exhibits an increased antagonistic activity with respect to a wide range of pathogenic and opportunistic pathogenic microorganisms and a resistance to quite a number of antibiotics.

Abstract: The present invention relates to methods of producing a polypeptide having biological activity in a bacterial cell, comprising: (a) cultivating a bacterial host cell in a medium conducive for production of the polypeptide, wherein the bacterial host cell comprises a nucleic acid construct comprising a promoter region operably linked to a polynucleotide sequence encoding the polypeptide and a modified mRNA processing/stabilizing sequence located downstream of the promoter region and upstream of the ribosome binding site of the polynucleotide sequence encoding the polypeptide, wherein the modified mRNA processing/stabilizing sequence promotes higher expression of the polynucleotide sequence compared to an unmodified mRNA processing/stabilizing sequence; and (b) isolating the polypeptide having biological activity from the cultivation medium.

Abstract: The invention provides a composition containing particulate composite of a polymer and a therapeutic agent. The composition also contains a complexing agent. The polymer interacts with the complexing agent in a host-guest or a guest-host interaction to form an inclusion complex. A therapeutic composition of the invention may be used to deliver the therapeutic agent and to treat various disorders. Both the polymer of the particulate composite and the complexing agent may be used to introduce functionality into the therapeutic composition. The invention also relates to a method of preparing a composition. The method combines a therapeutic agent, a polymer having host or guest functionality, and a complexing agent having guest or host functionality to form the therapeutic composition. The complexing agent forms an inclusion complex with the polymer. The invention also relates to a method of delivering a therapeutic agent.

Abstract: The present invention pertains to the discovery that B. anthracis possesses a luxS gene that encodes a functional LuxS polypeptide, and that B. anthracis synthesizes a functional AI-2 quorum-sensing molecule. The invention provides mutant B. anthracis bacteria lacking the function of the luxS gene, which do not produce a functional AI-2 molecule and have growth defects compared to wild-type B. anthracis. The invention also concerns methods for inhibiting the growth of B. anthracis, or for preventing or treating B. anthracis infection, by inhibiting the activity of the B. anthracis LuxS polypeptide, or by exposure of the B. anthracis to furanone. In particular, the invention concerns the use of furanone, a compound that inhibits AI-2-mediated quorum-sensing, to inhibit the growth of B. anthracis, to inhibit B. anthracis toxin production, particularly that of protective antigen, and to prevent or treat B. anthracis infection. The invention also provides methods to prevent B.

Abstract: The present invention relates to cells that have been genetically manipulated to have an altered capacity to express and/or produce proteins of interest. In particular, the present invention relates to modified host cells of Gram-positive microorganisms, such as Bacillus species that are capable of overexpressing ymaH. The invention encompasses polynucleotide constructs and expression vectors containing polynucleotide sequences that encode YmaH, and the modified host cells comprising them. In particular, the present invention relates to compositions and methods of overexpressing YmaH for enhancing the expression and production of proteins of interest (e.g., proteases) in Bacillus species.

Abstract: The invention relates to processes for producing an immunoglobulin or an immunologically functional immunoglobulin fragment containing at least the variable domains of the immunoglobulin heavy and light chains. The processes can use one or more vectors which produce both the heavy and light chains or fragments thereof in a single cell. The invention also relates to the vectors used to produce the immunoglobulin or fragment, and to cells transformed with the vectors.

Abstract: An object is to provide a means of highly producing an oxalate decarboxylase originating in a microorganism. A recombinant expression plasmid vector, which contains an ?-amylase promoter belonging to the genus Bacillus and an oxalate decarboxylase gene originating in a microorganism that is provided under the control of the promoter, is constructed. A host bacterium is transformed with this vector to prepare an oxalate decarboxylase producing bacterium. A recombinant oxalate decarboxylase is produced by culturing the producing bacterium and then recovering the oxalate decarboxylase thus produced.

Abstract: The present invention provides a mutant Bacillus bacterium capable of enhancing productivity of proteins or polypeptides, a recombinant microorganism produced by introducing genes encoding heterologous proteins or polypeptides into the mutant Bacillus bacterium, and a method for producing proteins or polypeptides by use of the recombinant microorganism. A mutant Bacillus bacterium which has, on the genome or plasmid thereof, DNA having a promoter sequence which is recognized and transcribed specifically during the sporulation stage and which is ligated to an upstream end of a sigA gene or a gene equivalent thereto, a recombinant microorganism produced by introducing genes encoding heterologous proteins or polypeptides into the mutant Bacillus bacterium, and a method for producing proteins or polypeptides by use of the recombinant microorganism.

Abstract: The present invention relates to methods of improving the introduction of DNA into bacterial host cells.

Abstract: The invention relates to a method for preparing closed bacterial ghosts by way of specific interactions between partners of a bioaffinity binding pair, and to the bacterial ghosts which can be obtained in this way. Active compounds can be packed into the closed bacterial ghosts. The closed ghosts can be employed in medicine, in the agricultural sphere and in biotechnology.

Abstract: The invention relates to a xylanase originating from a Bacillus strain. This xylanase is active over a wide range of acid and basic pH. The invention also relates to new strains of microorganisms producing this xylanase and to methods for preparing this xylanase. The invention also relates to a DNA molecule and to an expression vector or an integration vector containing this DNA molecule. The invention also relates to uses of the latter and to compositions containing it.

Abstract: The present invention provides a nucleic acid encoding nicotinamide phosphoribosyltransferase (NadV) from a V-factor independent bacterium and provides methods for using the gene as a selection marker for constructing recombinant bacteria from V-factor dependent bacteria. The method is an improvement over methods which rely on nucleic acids which confer antibiotic resistance for constructing recombinant bacteria. Methods for constructing attenuated recombinant Actinobacillus pleuropneumoniae using the selection method of the present invention are also provided.

Abstract: The present invention is directed to methods and compositions for microbial based production of pravastatin. The compositions of the invention include novel strains of microorganisms that are capable of efficiently hydroxylating compactin (ML-236 B) resulting in production of pravastatin. In particular, the microorganisms of the invention are genetically engineered to express both cytochrome P-450 and the fdxshe or fdxshe-like protein. The invention further relates to the use of such microorganisms in processes designed for production of pravastatin for use in treatment of disease such as hypercholesterolemia and hyperlipidemia.

Abstract: The present invention relates to tissue cement proteins produced by certain species of blood-feeding ectoparasites. These proteins and compositions comprising these proteins are particularly useful for the temporary or permanent bonding of animal tissues to each other or to other biomaterials. The present invention also relates to the use of tissue cement proteins in the production of vaccines that protect animals against the bite of blood-sucking ectoparasites and the transmission of viruses, bacteria and other pathogens by such ectoparasites.

Abstract: Novel insect inhibitory proteins are disclosed comprising two different components, both of which are required for biological activity. Various methods of linking both components together, so that a single protein provides insect inhibitory activity, are disclosed. Also disclosed are novel Bacillus thuringiensis nucleic acid sequences encoding Coleopteran-inhibitory crystal proteins, designated tIC100 (29-kDa) and tIC101 (14-kDa). Also disclosed are methods of making and using nucleic acid sequences in the development of the transgenic plant cells containing the novel nucleic acid sequences disclosed herein.

Abstract: The invention relates to an isolated and purified culture of Bacillus sp. strain 720/1 (LMG P-14798); to xylanases obtained from this strain and xylanases obtained from derivatives and mutants of strain 720/1. The invention also relates to a DNA molecule encoding a xylanase and to expression vectors or integration vectors containing the DNA molecule. This invention also relates to transformed host strains comprising the DNA molecule encoding the xylanase.

Abstract: The invention relates to a method for preparing closed bacterial ghosts by means of vesicle membrane fusion and to the bacterial ghosts which can be obtained in this way. Active compounds, e.g. genetic material, cell components, pharmaceutical and agricultural active compounds and also markers or dyes can be packaged in the closed bacterial ghosts. Metabolic functions and, where appropriate, the ability of the cells to proliferate can be restored on packaging genetic material in the bacterial ghosts. The closed ghosts can be used in medicine, in the agricultural sphere and in biotechnology.

Abstract: The invention relates to a method for increasing the copy number of a chromosomally integrated expression cassette in a microbial strain without leaving antibiotic resistance markers behind in the strain, the necessary genetic constructs, and the strains resulting from the method of the invention.

Abstract: Genes have been identified in the Bacillus genome that are responsive to various metabolic conditions and growth cycle changes. The new responsiveness of these genes allows for their use in regulated gene expression in Bacillus sp. and for the monitoring of bioreactor health.

Abstract: To provide a promoter, a recombinant DNA, a gene expression vector, an expression vector, and a transformant, which are capable of expressing a gene without inducing gene expression with an inducer; and a method for producing a protein and a kit therefor, which can be operated easily and performed inexpensively by convenient and inexpensive steps.

Abstract: The present invention provides for device for obtaining and storing genetic material. The device has a support and a head portion. The head portion is composed of a solid matrix for sorbing genetic material and a preserving mechanism for the protection of genetic material from degradation. Additionally, the present invention provides for a method of storing a sample of genetic material onto the swab device. In the preferred embodiment, the present invention utilizes polyester material as the solid matrix.

Abstract: The present invention provides a mutant aprE promoter and methods for the production of a desired protein in a Bacillus host cell, which comprises the mutant aprE promoter. The present invention provides the sequence of a preferred aprE mutant promoter, which provided for a 100-fold increase in the production of a protein from Bacillus subtilis.

Abstract: The invention is based on the discovery that recombinagenic oligonucleobases are active in prokaryotic cells that contain a strand transfer activity (RecA) and mismatch repair activity (MutS). Using this system a type of Duplex Mutational Vector termed a Heteroduplex Mutational Vector, was shown to be more active in prokaryotic cells than the types of mutational vectors heretofore tested. Further improvements in activity were obtained by replacing the tetrathymidine linker by a nuclease resistant oligonucleotide, such as tetra-2′-O-methyl-uridine, to link the two strands of the recombinagenic oligonucleobase and removing the DNA-containing intervening segment. The claims concern Duplex Mutational Vectors that contain the above improvements. In an alternative embodiment the claims concern the use of Duplex Mutational Vectors in prokaryotic cells.

Abstract: The present invention is directed to methods and compositions for TnpI-mediated genetic engineering using the Bacillus thuringiensis recombinase TnpI and the TnpI recombination substrates TRT or TRT′, and variants thereof. In particular, the invention relates to TRT or TRT′ sequences, and variants thereof, vectors, cells and kits useful for TnpI-mediated genetic recombination, as well as methods for the use of TRT or TRT′ sequences for TnpI-mediated genetic engineering.

Abstract: A nucleic acid derived from Bacillus thuringiensis contains a nucleotide sequence that encodes a polypeptide demonstrated to be toxic to fire ants.

Abstract: The invention concerns an in vitro process for altering the insect host range (spectrum) of pesticidal toxins. The process comprises recombining in vitro the variable region(s) (non-homologous) of two or more genes encoding a pesticidal toxin. Specifically exemplified is the recombining of the variable regions of two genes obtained from well-known strains of Bacillius thuringiensis var. kurstaki. The resulting products are chimeric toxins which are shown to have an expanded and/or amplified insect host range as compared to the parent toxins.

Abstract: A method for in vivo production of a library in cells comprising a multitude of mutated genetic elements, wherein an error-prone polymerase is used in each ancestral cell to replicate all or a part of a genetic element independently of the host chromosomal replication machinery. The genetic element comprises i) an origin of replication from which replication is initiated, ii) optionally a genetic marker, e.g. a gene conferring resistance towards an antibiotic, iii) a gene encoding the polypeptide of interest. Also methods for the generation of a DNA sequence encoding a desired variant of a polypeptide of interest, and for the determination of such a DNA sequence are described.

Abstract: The present invention provides methods for building DNA constructs in vitro, transforming such constructs into competent Bacillus strains with good efficiency, and generating populations of mutants. Also provided is a method to assemble DNA constructs in situ.

Abstract: The present invention provides a isolated bacteriophage useful as a tool for studying biological, biochemical, physiological and genetic properties of actinomycetes and other organisms which comprises a novel strain of Saccharomonospora having certain specified characteristics. The invention also relates to a process for the isolation of the said bacteriophage and/or DNA phage and to a novel universal growth medium which is particularly useful in the said process. Another embodiment of the process relates to a cloning vector which comprises a plasmid or bacteriophage comprising the phage DNA of the invention.

Abstract: Genes have been identified in the Bacillus genome that are responsive to various metabolic conditions and growth cycle changes. The new responsiveness of these genes allows for their use in regulated gene expression in Bacillus sp. and for the monitoring of bioreactor health.

Abstract: Disclosed herein are novel phenol oxidizing enzymes naturally-produced by strains of the species Stachybotrys which possess a pH optima in the alkaline range and which are useful in modifying the color associated with dyes and colored compounds, as well as in anti-dye transfer applications. Also disclosed herein are biologically-pure cultures of strains of the genus Stachybotrys, designated herein Stachybotrys parvispora MUCL 38996 and Stachybotrys chartarum MUCL 38898, which are capable of naturally-producing the novel phenol oxidizing enzymes. Disclosed herein is the amino acid and nucleic acid sequence for Stachybotrys phenol oxidizing enzymes as well as expression vectors and host cells comprising the nucleic acid. Disclosed herein are methods for producing the phenol oxidizing enzyme as well as methods for constructing expression hosts.

Abstract: A host cell which is provided with a S-layer comprising a fusion polypeptide consisting essentially of:

Abstract: This invention relates to ethanol production as a product of bacterial fermentation.

Abstract: The invention relates to processes for producing an immunoglobulin or an immunologically functional immunoglobulin fragment containing at least the variable domains of the immunoglobulin heavy and light chains. The processes can use one or more vectors which produce both the heavy and light chains or fragments thereof in a single cell. The invention also relates to the vectors used to produce the immunoglobulin or fragment, and to cells transformed with the vectors.

Abstract: The invention relates to a method for producing an integrant(s) of Bacillus thuringiensis which produces a larger quantity of a crystal delta-endotoxin with greater pesticidal activity as compared to the crystal delta-endotoxin produced by the corresponding parental strain. The crystal delta-endotoxin produced by the integrant Bacillus thuringiensis will have an activity directed towards the same pest(s) as its parent Bacillus thuringiensis crystal delta-endotoxin. The invention further relates to such integrants, compositions comprising such integrants, as well as methods for controlling a pest(s) using these compositions.

Abstract: Novel alpha-amylase mutants derived from the DNA sequences of naturally occurring or recombinant alpha-amylases are disclosed. The mutant alpha-amylases, in general, are obtained by in vitro modifications of a precursor DNA sequence encoding the naturally occurring or recombinant alpha-amylase to generate the substitution (replacement) or deletion of one or more oxidizable amino acid residues in the amino acid sequence of a precursor alpha-amylase. Such mutant alpha-amylases have altered oxidative stability and/or altered pH performance profiles and/or altered thermal stability as compared to the precursor. Also disclosed are detergent and starch liquefaction compositions comprising the mutant amylases, as well as methods of using the mutant amylases.

Abstract: Methods of preparing recombinant Bacillus anthracis protective antigen or a variant or fragment thereof for use in vaccines is disclosed. The protein is expressed in a recombinant microorganism which comprises a sequence which encodes PA or said variant or fragment thereof wherein either (i) a gene of the microorganism which encodes a catabolic repressor protein and/or AbrB is inactivated, and/or (ii) wherein a region of the PA sequence which can act as a catabolic repressor binding site and/or an AbrB binding site is inactivated. Useful quantities of protein are obtainable from these organisms.

Abstract: A composition for expression of a protein-encoding gene in a host cell is described, making use of an heterologous regulon. This composition provides a protein-encoding gene under control of a promoter heterologous to the host cell, and a gene for an RNA polymerase, preferably a single-subunit RNA polymerase that recognizes the promoter heterologous to the host cell. The gene for the RNA polymerase is under control of an inducible promoter recognized by the host cell. Also disclosed is a method for expressing the protein-encoding gene using the composition described.

Abstract: This invention relates to an isolated nucleic acid fragment encoding a glycolysis or respiration protein. The invention also relates to the construction of a chimeric gene encoding all or a portion of the glycolysis or respiration protein, in sense or antisense orientation, wherein expression of the chimeric gene results in production of altered levels of the glycolysis or respiration protein in a transformed host cell.

Abstract: Novel methods and novel industrial unicellular microorganism strains, particularly industrial Bacillus strains, are provided for enhanced production of endogenous and exogenous polypeptides. Cloning vehicles containing the gene expressing the polypeptide of interest are introduced into a compatible host. Transformed hosts harboring the introduced vehicle in a stable way by integration of the vehicle into the host cells chromosome are selected. Efficient transfer of the vehicle containing the gene of interest is achieved, with the resulting industrial strain transformants being effective, stable producers of the desired polypeptide product.

Abstract: The present invention discloses a selection marker free system which can be used to introduce genetic modifications in bacteria, yeasts and fungi. The system can be employed to introduce or delete desired genes or DNA fragments in the genome of the indicated host species without leaving any undesired DNA i.e. the selection marker used for selection of transformants or other DNA used for cloning. In this way strains have been developed containing only desired genes introduced at desired chromosomal sites. Similarly, desired DNA fragments have been deleted or replaced at desired sites.

Abstract: A positive selection vector is provided for the transformation and screening of Gram positive bacteria and particularly Bacillus sp. for the presence of foreign DNA. The vector comprises a mutant gene encoding a signal peptide processing mutation. Expression of the mutant gene in a Bacillus host cell which lacks the ability to metabolize sucrose is lethal when cells are grown in the presence of sucrose. Foreign DNA may be inserted into the vector so as to inactivate the mutant gene thereby permitting the cells to grow in the presence of sucrose and allowing for facile selection of transformants.

Abstract: The present invention relates to methods of producing a polypeptide, comprising: (a) cultivating a mutant of a Bacillus cell, wherein the mutant (i) comprises a first nucleic acid sequence encoding the polypeptide and a second nucleic acid sequence comprising a modification of at least one of the genes responsible for the biosynthesis or secretion of a surfactin or isoform thereof under conditions conducive for the production of the polypeptide and (ii) the mutant produces less of the surfactin or isoform thereof than the Bacillus cell when cultured under the same conditions; and (b) isolating the polypeptide from the cultivation medium. The present invention also relates to mutants of Bacillus cells and methods for producing the mutants.

Abstract: The invention relates to a method for producing an integrant(s) of Bacillus thuringiensis. The invention further relates to such integrants, compositions comprising such integrants, as well as methods for controlling a pest(s) using these compositions.

Abstract: Increased production of a heterologous cellulase is achieved by transforming Bacillus subtilis and Bacillus licheniformis with genetic constructs containing a Bacillus licheniformis ATCC 53926 protease promoter and signal sequence to express alkalophilic cellulase genes.