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: Methods and devices are provided for gene therapy using encapsulated packaging cell lines to deliver viral particles carrying at least one heterologous gene encoding at least one biologically active molecule.

Abstract: Described are preferred myocardial grafts of skeletal myoblasts or cardiomyocytes, and cellular compositions and methods useful in obtaining the grafts. The myocardial grafts are stable and can be used, for example, to deliver recombinant proteins directly to the heart.

Abstract: An objective of the present invention is to provide a packaging cell for preparing a retrovirus having a high viral titer. The cell producing a recombinant retrovirus is constructed by introducing, the Polyoma virus early region gene together with a recombinant plasmid or a recombinant retrovirus free from any replication origin derived from Polyoma virus into a packaging cell for preparing a recombinant retrovirus.

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: Lentiviral vectors modified at the 5' LTR or both the 5' and 3' LTR's are useful in the production of recombinant lentivirus vectors. Such vectors can be produced in the absence of a functional tat gene. Multiple transformation of the host cell with the vector carrying the transgene enhances virus production.

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: The present invention provides a system for controlled transgene transcription using chimeric activator and repressor proteins functioning in a novel regulatory network. The target transgene is actively silenced in non-inducing conditions by a novel class of chimeric proteins consisting of the DNA-binding tetracycline repressor fused to distinct multimerized eukaryotic transcriptional repression domains. In the presence of a tetracycline "inducer", the repressor does not bind to the promoters for both the target gene and for another regulatory gene encoding a transactivator (e.g., GAL4-VP16). Under these inducing conditions, the transactivator activates expression of the target transgene and of its own gene, in an additional autoregulatory positive feedback loop.

Abstract: The invention relates to modified two-hybrid systems, in particular a reverse two-hybrid system which employs as a reporter a gene encoding a modifying agent such as an enzyme, and a signal agent which is modified by the enzyme usually by being broken down, such that in the event of an inhibition of binding of the two hybrid proteins a detectable signal is produced. The system is particularly useful for drug screening.

Abstract: Methods and a kit are provided for characterizing small molecules from a library of small molecules or alternatively identifying protein targets to which known small molecules bind. The methods include forming hybrid ligand in which at least one ligand is a small molecule. The hybrid ligand is introduced into cells that in turn contain a first and a second expression vector. Each expression vector includes DNA for expressing a hybrid protein that encodes a target protein linked to a coding sequence for a transcriptional module. The cells further contains a reporter gene, the expression of which is conditioned on the proximity of the first and second hybrid proteins to each other, an event that occurs only if the hybrid ligand binds to target sites on both hybrid proteins. Those cell which express the reporter gene are selected and the unknown small molecule or the unknown hybrid protein is identified.

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: Transformed cell lines containing a reporter gene operatively linked to a genetic control element that is responsive to growth factor-stimulated cell proliferation and/or oncogene-mediated neoplastic transformation are provided. Also provided are methods for using such transformed cell lines to screen for growth factor antagonists and/or antineoplastic agents.