Abstract: The present invention provides mouse models of growth hormone insensitivity including Laron syndrome. In particular, the present invention provides transgenic mice incapable of expressing functional growth hormone receptor including mice which further cannot express functional growth hormone binding protein. The invention further provides methods for testing the usefulness of chemical compounds in the treatment of growth hormone insensitivity and diabetic end-organ disease.

Abstract: Transgenic mice with a non-functional proteinase activated receptor-2 (PAR2) gene are prepared by targeted disruption of the endogenous PAR2 gene. The resulting transgenic mice display a phenotype including a lack of a hypotensive response to administration of the peptide, SLIGRL, and a reduction in carrageenin-induced paw edema compared to wild type mice.

Abstract: The present invention relates to a method of gene or DNA targeting in cells of vertebrate, particularly mammalian, origin. That is, it relates to a method of introducing DNA into primary or secondary cells of vertebrate origin through homologous recombination or targeting of the DNA, which is introduced into genomic DNA of the primary or secondary cells at a preselected site. The present invention further relates to primary or secondary cells, referred to as homologously recombinant (HR) primary or secondary cells, produced by the present method and to uses of the homologously recombinant primary or secondary cells. The present invention also relates to a method of turning on a gene present in primary cells, secondary cells or immortalized cells of vertebrate origin, which is normally not expressed in the cells or is not expressed at significant levels in the cells.

Abstract: The present invention discloses MmRad51-deficient transgenic mice and mouse cells, as well as MmRad51/p53-deficient transgenic mice and mouse cells. Also described is a method of screening for proteins that rescue the senescence phenotype in MmRad51/p53-deficient cells.

Abstract: The present invention provides mice which are deficient in the normal expression of one or more members of the RAR or RXR class of receptors, to mice heterozygous for such deficiency, and to cell lines, preferably pluripotent or totipotent cell lines, which are heterozygous or homozygous for such deficiency. The present invention further provides the use of any of the above mice and cell lines in situations where the absence of at least one RAR or RXR receptors, or the normal expression thereof, is desirable.

Abstract: The invention provides a nonhuman transgenic animal having a transgene disrupting or interfering with expression of G.alpha..sub.q chromosomally integrated into the germ cells of the animal. Cells and cell lines derived from these nonhuman transgenic animals; a method for producing a transgenic nonhuman animal having a phenotype characterized by expression of a transgene which is otherwise not naturally occurring, where the expression of the transgene disrupts or interferes with G.alpha..sub.q activity; a method for determining an effect of an agent on G.alpha..sub.q expression, by administering an effective amount of the agent to a transgenic nonhuman animal having a transgene disrupting or interfering with G.alpha..sub.q, and measuring a physiological response of the transgenic nonhuman animal to the agent, and comparing the physiological response of the transgenic nonhuman animal having a transgene disrupting or interfering with G.alpha..sub.q to a control animal is also provided.

Abstract: Selectable Herpesvirus saimiri vectors which have a selection gene inserted into a junction region of the L- and H-DNA are described. Vectors of this type are able persistently to infect human T cells and thus are suitable for the expression of foreign genes in human T cells. An additional advantage is that no infectious virus particles are produced during this.

Abstract: Provided are transgenic mice genetically engineered for a deficiency of the heart-skeletal muscle isoform of the adenine nucleotide translocator protein (Ant1). These mice exhibit histological, biochemical and physiological signs of deficiency in oxidative phosphorylation and energy generation, and these mice provide the first animal model for mitochondrial myopathy and hypertrophic cardiomyopathy. This animal model is used in methods for testing compounds for therapeutic value in treating failure to exchange ATP and ADP across the mitochondrial inner membrane, OXPHOS deficiency and in treating cardiac hypertrophy.

Abstract: Chimeric proteins are provided comprising a first domain that attaches said chimeric protein to a target nucleic acid, and a second domain that integrates donor nucleic acid into a target nucleic acid. Also provided are nucleic acid constructs, recombinant vectors encoding invention chimeric proteins, recombinant retroviruses, and related methods.

Abstract: A method for gene therapy using small fragment homologous replacement. The method introduces small fragments of exogenous DNA into regions of endogenous genomic DNA virtually homologous to the exogenous DNA. The exogenous DNA fragments contains sequence modification that correct mutations in the endogenous DNA or introduce mutations that alter cellular or an infecting pathogen phenotype.

Abstract: A transgenic mouse whose genome comprises a disruption of the endogenous growth differentiation factor-11 (GDF-11) gene is disclosed. Also disclosed are methods for making such mice. The mice exhibit a phenotype of increased muscle tissue.

Abstract: The invention is based on the discovery that Duplex Mutational Vectors 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 Non-Chimeric Mutational Vector, having no RNA:DNA hybrid-duplex, was shown to be active in prokaryotic cells if protected from 3' exonuclease attack. Such protection can be conferred by replacement of a tetrathymidine linker by a nuclease resistant oligonucleotide, such as tetra-2'-O-methyl-uridine, to link the two strands of the recombinagenic oligonucleobase. The claims concern 3'-exonuclease protected Non-Chimeric Mutational Vectors.

Abstract: A transgenic mouse with alterations in the H2-Ma gene is prepared by introduction of an altered H2-Ma gene into a host mouse. The resulting transgenic mice do not produce functional H2-M molecules.

Abstract: The method of the invention provides the use of a replacement-type targeting construct to delete large fragments of genomic DNA by gene targeting. The replacement targeting construct, which may contain a selectable marker, is constructed to contain two regions of sequences which are homologous to the 5' and 3' flanking sequences of the targeted locus. After transfection of the targeting construct into the desired cell line, gene targeted-mediated deletions are identified by selection and further characterized. The invention is useful in any situation where one would want to create a large genomic deletion. Examples of suitable loci include MHC Class I and II antigens and immunoglobulin genes, including, for example, variable and constant region of kappa, lambda, or heavy chains.

Abstract: The present invention provides a method for preparing HSV vectors. The method comprises co-transfecting a source vector and a mutating cassette together into a population of appropriate host cells, such that homologous recombination occurs between the mutating cassette and the source vector whereby the mutating cassette replaces a region of the HSV genome. The mutating cassette has a unique restriction site not present in the sequence of the vector. The method further comprises plaquing the co-transfected host cells, selecting plaques in which recombination has occurred between the source vector and the mutating cassette, and isolating the viral DNA from the plaques. The isolated viral DNA is digested with a restriction endonuclease appropriate for cleaving the viral DNA at the unique restriction site within the mutating cassette to produce two viral polynucleotides. Following purification, the two viral polynucleotides can be ligated to form an HSV vector comprising the two viral polynucleotides.

Abstract: A method for achieving site specific integration of a desired DNA at a target site in a mammalian cell via homologous recombination is described. This method provides for the reproducible selection of cell lines wherein a desired DNA is integrated at a predetermined transcriptionally active site previously marked with a marker plasmid. The method is particularly suitable for the production of mammalian cell lines which secrete mammalian proteins at high levels, in particular immunoglobulins. Novel vectors and vector combinations for use in the subject cloning method are also provided.

Abstract: A transgenic mouse whose genome comprises a disruption of the endogenous growth differentiation factor-8 (GDF-8) gene is disclosed. Also disclosed are methods for making such mice. The transgenic mice exhibit a phenotype of increased muscle tissue.

Abstract: Disclosed is a method for the production of deletions in the chromosomal DNA of a single eukaryotic cell. More specifically, the method involves the creation of either random chromosomal deletions, or chromosomal deletions within a predetermined genetic loci. The deletions are created by integration of a DNA construct into the chromosomal DNA of the single eukaryotic cell, followed by spontaneous or irradiation induced deletion of the DNA construct from the chromosomal DNA. Also, disclosed is a method for the production of deletions in the chromosomal DNA of a multicell organism using a DNA construct.

Abstract: Methods and compositions are provided for transferring DNA sequence information from a first vector to a second vector. In the subject methods, a first vector comprising a region of DNA having a sequence of interest is contacted with a set of three pairs of oligonucleotide primers under conditions sufficient to produce three different PCR products, where each product corresponds to a different reading frame. The oligonucleotide primers comprise a first region of sequence identity with the first vector and a second region permissive of site specific recombination with the second vector. The resultant PCR products are combined with the second vector under conditions sufficient for site specific recombination to occur. Also provided are kits for use in performing the subject methods.

Abstract: Switch regions derived from an immunoglobulin (Ig) gene are used to direct recombination between a targeting construct containing a promoter, a switch region (S.sub.1), and 2) a target locus minimally containing a promoter, a switch region (S.sub.2), and a target sequence.

Abstract: The invention relates to sucrose isomerases, to DNA sequences coding therefor, and to novel processes for producing noncariogenic sugars.

Abstract: Methods and compositions are provided for expression of mammalian genes in culture. An amplifiable gene is introduced by homologous recombination in juxtaposition to a target gene, the resulting combination of amplifiable gene and target gene transferred to a convenient host and the target gene amplified by means of the amplifiable gene. The resulting expression host may then be grown in culture with enhanced expression of the target gene.

Abstract: The present invention provides a transgenic mouse comprising a disrupted hepsin gene. In particular, the invention provides methods of making the transgenic mouse comprising the disrupted hepsin gene by utilizing a hepsin targeting vector for homologous recombination in mouse embryonic stem cells. Also, nucleotide and amino acid hepsin sequences are disclosed.

Abstract: A system is described which utilizes a novel system of repressor titration for maintenance of a plasmid useful in gene therapy and production of a recombinant protein. The system utilizes a transformed host cell containing a plasmid including an operator susceptible to binding by a repressor expressed in trans, a first chromosomal gene encoding the repressor, and a second chromosomal gene that is functionally associated with an operator and essential for cell growth, wherein the plasmid is present in the cell in sufficient numbers to titrate the repressor such that the essential gene is expressed, thereby permitting cell growth.

Abstract: A transgenic animal with alterations in an thrombin receptor gene is prepared by introduction of an altered thrombin receptor gene into a host animal. The resulting transgenic animals do not produce functional thrombin receptor molecules.

Abstract: A recombinant vector useful for regulating the expression of a gene contred by a baculovirus polyhedrin promoter or P10 promoter, via a sequence forming an RAR-type hormone receptor binding site.

Abstract: The subject invention pertains to novel Entomopoxvirus (EPV) polynucleotide sequences free from association with other viral sequences with which they are naturally associated, recombinant polynucleotide vectors containing the sequences, recombinant viruses containing the sequences, and host cells infected with the recombinant viruses are provided herein, as well as methods for use thereof in the expression of heterologous proteins in both insect and mammalian host cells.

Abstract: Disclosed are (1) a DNA sequence encoding a mutant L3T4 protein which, when expressed on the surface of a cell, is capable of facilitating infection of the cell by human immunodeficiency virus; the mutant protein includes at least one amino acid residue substitution or deletion in a segment corresponding to the gp120 binding epitope of a native L3T4 protein so as to increase homology between that segment and its counterpart in a CD4 protein; (2) a murine cell line or strain transfected with such a DNA sequence; and (3) a transgenic mouse susceptible to infection by human immunodeficiency virus.

Abstract: The invention provides for a methods and compositions for identifying proteins or compounds that directly or indirectly modulate a genomic polynucleotide and methods for identifying active genomic polynucleotides. Generally, the method comprises inserting a BL (beta-lactamase) expression construct into an eukaryotic genome, usually non-yeast, contained in at least one living cell, contacting the cell with a predetermined concentration of a modulator, and detecting BL activity in the cell.

Abstract: Transgenic mice carrying two transgenes, the first coding for a transactivator fusion protein comprising a tet repressor and a polypeptide which directly or indirectly activates in eucaryotic cells, and the second comprising a gene operably linked to a minimal promoter operably linked to at least one tet operator sequence, are disclosed. Isolated DNA molecules (e.g., targeting vectors) for integrating a polynucleotide sequence encoding a transactivator of the invention at a predetermined location within a second target DNA molecule by homologous recombination are also disclosed. Transgenic mice having the DNA molecules of the invention integrated at a predetermined location in a chromosome by homologous recombination are also encompassed by the invention. Methods to regulate the expression of a tet operator linked-gene of interest by administering tetracycline or a tetracycline analogue to a mouse of the invention are also disclosed.

Abstract: The present invention relates to transgenic mice in which the biological function of at least one cell cycle regulatory proteins of the INK4 family is altered.

Abstract: This invention is directed toward the characterization and cloning of a cAMP-responsive transcription enhancer binding protein (CREB). This protein, CREB, is a transcriptional activator which activates transcription in eukaryotic cells. This CREB protein can be used to increase or decrease production of proteins by stimulating expression of a recombinant gene that is operably-linked to the CRE enhancer element and responsive to cAMP.

Abstract: The present invention relates to a tetracycline repressor-mediated binary regulation system for the control of gene expression in transgenic mice. It is based, at least in part, on the discovery that, in a transgenic mouse that carries a first transgene under the control of a modified promoter comprising a tetR operator sequence and a second transgene encoding the tetR protein, expression of the first transgene may be efficiently induced by administering tetracycline to the mouse.

Abstract: The invention described here is a method whereby a molecular tag is put on a gene, transcript and protein in a single recombinational event. The protein tag takes the form of a unique peptide that can be recognized by an antibody or other specific reagent, the transcript tag takes the form of the sequence of nucleotides encoding the peptide that can be recognized by a specific polynucleotide probe, and the gene tag takes the form of a larger sequence of nucleotides that includes the peptide-encoding sequence and other associated nucleotide sequences. The central feature of the invention in its essential form is that the tag-creating DNA has a structure such that when it is inserted into an intron within a gene it creates two hybrid introns separated by a new exon encoding the protein tag. A major virtue of the method is that it allows one to identify new proteins or protein-containing structures, and, having done so, to readily identify and analyze the genes encoding those proteins.

Abstract: The present invention relates to a method for the controlled expression, in a lactic acid bacterium, of a DNA fragment containing one or more genes coding for a desired characteristic, wherein the DNA fragment is under the control of a promoter for a microbial gene which codes for an antimicrobial peptide, and the gene or genes are brought to expression on the DNA fragment by the addition of a suitable inducing factor for the transcription activation. The promoter is preferably the nisA promoter from Lactococcus lactis. The inducer is preferably acceptable for food products, and more preferentially is nisin or derivatives thereof. The invention also relates to a method for the production of proteins or RNA, as well as to a method for the preparation of dairy products using the expression system according to the invention. The invention finally relates to lactic acid bacteria and expression vectors for use in the method according to the invention.

Abstract: The present invention provides an isolated nucleic acid sequence encoding murine IL-2R.gamma.. The present invention also provides a vector comprising a mutated IL-2R.gamma. nucleic acid which is capable of homologous recombination in at least some cells to which the vector is introduced. The present invention also provides an embryonic stem cell comprising a mutated IL-2R.gamma. nucleic acid integrated into the cell by homologous recombination following transfection with the vector above. The present invention further provides a blastocyst cell comprising the embryonic stem cell above. In addition, the present invention provides a transgenic animal comprising a mutated IL-2R.gamma. gene. In particular, the animal is a non-human mammal whose germ and somatic cells contain a mutated IL-2R.gamma. gene sequence introduced into said mammal, or an ancestor thereof, at an embryonic stage.

Abstract: The present invention relates to methods of producing a polypeptide, comprising: (a) introducing into a respiratory-defective mutant of a cell (i) one or more first nucleic acid sequences which complement the respiratory defect and (ii) a second nucleic acid sequence which encodes the polypeptide; (b) cultivating the cell containing the first and second nucleic acid sequences in a culture medium under aerobic conditions suitable for expression of the first and second nucleic acid sequences; and (c) isolating the polypeptide from the cultivation medium of the cell. The present invention also relates to methods for disrupting a gene in a respiratory-deficient mutant cell. The present invention further relates to respiratory-deficient mutant cells and methods for obtaining such mutant cells.

Abstract: Disclosed are compositions and methods for inhibiting hexokinase enzymes in mammalian cells. Specifically provided are proteins that stimulate the production of trehalose-6-phosphate and their respective genes; hexokinase-specific ribozymes and genes encoding such constructs; and agents that competitively reduce hexokinase activity, e.g., by displacing hexokinase from mitochondria, and their respective genes. The latter group of agents includes inactive hexokinases and fragments thereof that retain mitochondrial binding functions and hexokinase-glucokinase chimeras that further substitute glucokinase activity for hexokinase activity. Mammalian cells including such hexokinase inhibitors, methods of making such cells and various in vitro and in vivo methods of using cells with reduced hexokinase activity are also described herein.

Abstract: A gene activation/inactivation and site-specific integration system has been developed for mammalian cells. The invention system is based on the recombination of transfected sequences by FLP, a recombinase derived from Saccharomyces. In several cell lines, FLP has been shown to rapidly and precisely recombine copies of its specific target sequence. For example, a chromosomally integrated, silent .beta.-galactosidase reporter gene was activated for expression by FLP-mediated removal of intervening sequences to generate clones of marked cells. Alternatively, the reverse reaction can be used to target transfected DNA to specific chromosomal sites. These results demonstrate that FLP can be used, for example, to mosaically activate or inactivate transgenes for a variety of therapeutic purposes, as well as for analysis of vertebriate development.

Abstract: The invention provides methods and compositions for evaluating modulators of the Stat6 signaling pathway; in a particular, transgenic mice comprising a transgene within a Stat6 allele locus, encoding a selectable marker and displacing the SH2-encoding domain of the Stat6 allele. More particularly, the transgene may comprise 3' and 5' regions with sufficient complementarity to the natural Stat6 allele at the locus to effect homologous recombination of the transgene with the Stat6 allele. Such mice provide useful animal models for determining the effect of candidate drugs on a host deficient in Stat6 function. The invention provides a variety of methods for making and using the subject compositions; in particular, methods for determining the effect of a candidate drug on a mouse deficient in Stat6 function and methods of evaluating the side effects of a Stat6 function inhibitor.

Abstract: A production process of optically active amino acids comprising causing a microorganism belonging to Rhodococcus, Mycobacterium, Arthrobacter, Nocardiopsis or Bacillus sp. and having nitrile-hydrolyzing activity to act on a nitrile or derivative thereof.

Abstract: A bar code detection circuit accepts as input the discretized analog output of a CCD array, and performs piecewise linear reconstruction to produces a continuous polylinear output signal. In the region of a bar/space transition, the output signal is a close approximation of the reflectance function of a bar code symbol convolved with the system transfer function of the bar code reader. Linear interpolation is performed in order to determine the offset of a given threshold value from an edge of the CCD analog output.