Abstract: This invention relates to delivery of biologically active cargo molecules, such as polypeptides and nucleic acids, into the cytoplasm and nuclei of cells in vitro and in vivo. Intracellular delivery of cargo molecules according to this invention is accomplished by the use of novel transport polypeptides which comprise HIV tat protein or one or more portions thereof, and which are covalently attached to cargo molecules. The transport polypeptides in preferred embodiments of this invention are characterized by the presence of the tat basic region (amino acids 49-57), the absence of the tat cysteine-rich region (amino acids 22-36) and the absence of the tat exon 2-encoded carboxy-terminal domain (amino acids 73-86) of the naturally-occurring tat protein. By virtue of the absence of the cysteine-rich region, the preferred transport polypeptides of this invention solve the potential problems of spurious trans-activation and disulfide aggregation.

Abstract: Substantially pure single-stranded oligonucleotides having a preselected sequence of not more than about 200 nucleotides, at least one of which is at a preselected position in the sequence and includes a base with a covalently attached linker arm containing or capable of binding at least one reporter group or solid support. A process for the chemical synthesis of the substantially pure single-stranded oligonucleotide and modified nucleosides useful in such synthesis are provided.

Abstract: Disclosed are oligonucleotide analogs comprising oligonucleoside sequences having from 3 to about 200 bases and containing internucleoside linkages wherein amide linkages replace phosphodiester linkages that are the backbones of the natural oligonucleotides that make up RNA and DNA. Also disclosed are bifunctional nucleoside analogs, a process for preparing dimers and trimers therefrom, and a method of using these bifunctional nucleoside intermediates, including the dimers and trimers, to synthesize the above-described oligonucleotide analogs using conventional synthetic organic procedures known in the art, preferably in a solid phase synthesis, more preferably in an automated peptide synthesizer.

Abstract: Oligonucleotides having the general formula, ##STR1## in which B denotes an arbitrary nucleobase,A equals 0 or CH.sub.2 ;X or Z equals O, S, NH or denotes CH.sub.2 whereby X and Z can be the same or different,V and W denote O, S, Se, NH.sub.2 or an alkyloxy residue, or OH or SH whereby V and W can be the same or different in a monomer unit andL is a H atom or a partner of a binding pair andC equals --O--R and R is an alkyl group with at least 1 C atom which may be modified if desired, or it denotes an alkenyl or alkynyl group with at least 2 C atoms which may be modified if desired, whereby the modification consists of a substitution by one or several halogen, cyano, carboxy, hydroxy, nitro or/and mercapto residues, andn is an arbitrary whole number, are stable antisense probes which bind specifically. Such oligonucleotides and polynucleotides may be used for the regulation of gene expression and as pharmaceutical agents.

Abstract: Oligonucleotides are provided which are targeted to nucleic acids encoding human c-raf and capable of inhibiting raf expression. The oligonucleotides may have chemical modifications at one or more positions and may be chimeric oligonucleotides. Methods of inhibiting the expression of human raf using oligonucleotides of the invention are also provided. The present invention further comprises methods of inhibiting hyperproliferation of cells and methods of treating abnormal proliferative conditions which employ oligonucleotides of the invention.

Abstract: The present invention provides a nucleotide-immobilized support which includes an insoluble support and a polynucleotide bound to the support. This polynucleotide includes at least one sequence complementary to a polyadenylic acid tail of a mRNA. The nucleotide-immobilized support is useful for a variety of methods, including the synthesis of both sense and antisense cDNA, as well as double stranded cDNA. The nucleotide-immobilized support is also useful for synthesis of both sense and antisense mRNA. Additionally, the nucleotide-immobilized support also provides an improved method of placing a cDNA sequence into a vector in a particular orientation. The present invention also includes a new method for binding polynucleotides to insoluble supports.

Abstract: This invention relates to delivery of biologically active cargo molecules, such as polypeptides and nucleic acids, into the cytoplasm and nuclei of cells in vitro and in vivo. Intracellular delivery of cargo molecules according to this invention is accomplished by the use of novel transport polypeptides which comprise HIV tat protein or one or more portions thereof, and which are covalently attached to cargo molecules. The transport polypeptides in preferred embodiments of this invention are characterized by the presence of the tat basic region (amino acids 49-57), the absence of the tat cysteine-rich region (amino acids 22-36) and the absence of the tat exon 2-encoded carboxy-terminal domain (amino acids 73-86) of the naturally-occurring tat protein. By virtue of the absence of the cysteine-rich region, the preferred transport polypeptides of this invention solve the potential problems of spurious trans-activation and disulfide aggregation.

Abstract: Disclosed are ribozyme analogs having the ability to endonucleolytically cleave a sequence of 3' to 5' linked ribonucleotides. The ribozyme analogs include a plurality of 3' to 5' covalently-linked nucleotides, and a rigid molecular linker having at least one non-nucleotidic molecule covalently linked to two of the nucleotides. Also disclosed are methods of preparing and utilizing the ribozyme analogs of the invention, and pharmaceutical formulations and kits containing such ribozyme analogs.

Abstract: Chemical compounds consisting of an alpha or beta oligo-4'-thioribonucleotide or oligo-4'-thio-2'-deoxyribonucleotide characterized in that they comprise a concatenation of 4'-thioribonucleotides or 4'-thio-2'deoxyribonucleotides, respectively. The concatenation is optionally linked to an effector, in particular a radical corresponding to an intercalating agent or a photoactivatable or chemical radical, e.g. a radical carrying a function which reacts directly or indirectly with the nucleotide chains, or a radical whose presence enables easy and sensitive detection. Methods for preparing said compounds, and their uses in therapeutics, diagnostics and as laboratory reagents, are also described.

Abstract: Compounds are provided having the structure: ##STR1## wherein the L groups are spanner or linker units, the Y and T group are functional groups for interacting with target molecules of interest, the X groups are oxygen or sulfur and the E groups are H, conjugate groups or intermediate groups used during the synthesis of the compounds are prepared using H phosphonate type chemistry wherein the functional groups are added during an oxidization step.

Abstract: Ribozymes designed to provide improved rates of catalytic turnover are described. The compounds of this invention comprise a catalytic region, at least one substrate binding region, and at least one displaceable antisense arm, whereby the rate of release of the endonuclease cleavage fragments is enhanced. A method to make such ribozymes is also described.

Abstract: Oligonucleotide-mimicking macromolecules that have improved nuclease resistance are provided. Replacement of the normal phosphorodiester inter-sugar linkages found in natural oligonucleotides with three or four atom linking groups provide unique oligonucleotide-mimicking macromolecules that are useful in regulating RNA expression and in therapeutics. Methods of synthesis and use also are disclosed.

Abstract: Tetrahydrofuranyl compounds are provided that are functionalized to include pendant conjugate groups, and which are useful in diagnostic assays and as research reagents. Novel intermediates for the synthesis of the compounds are also provided.

Abstract: The present invention relates to a DNA fragment which contains a gene responsible for the function of autonomous replication of plasmid in a Coryneform bacterium, said gene being obtained from plasmid pBY503 held in Brevibacterium stationis, and in which at least one point mutation capable of increasing the copy number of plasmid exists on said gene region. By using a Coryneform transformed with the vector constructed using said DNA fragment and an industrially useful gene such as aspartase gene or tryptophan synthase gene, production of the useful product occurs with higher efficiency than conventional methods.

Abstract: The invention concerns a complex between at least one negatively charged nucleic acid and at least one positively charged polymeric conjugate, the link between the nucleic acid and the polymeric conjugate being electrostatic in nature, the polymeric conjugate containing a polymer formed from monomer components having free NH.sub.3.sup.+ functions of the aforementioned components and being as follows:--the free NH.sub.3.sup.+ functions from the aforementioned components are substituted in a ratio of at least 10%, advantageously from 45% to 70%, particularly 60%, by noncharged residues leading to a reduction of positive charges in comparison to the same nonsubstituted polymeric conjugate, facilitating the release of nucleic acid by the dissociation of the complex,--the aforementioned residues possess in addition the following properties:.fwdarw.they contain at least one hydroxyl group,.fwdarw.they do not correspond to a recognition signal recognized by a cellular membrane receptor,--the free NH.sub.3.sup.

Abstract: Novel oligomers are disclosed which have enhanced ability with respect to forming duplexes or triplexes compared with oligomers containing only conventional bases. The oligomers contain 7-deaza-7-substituted purines or related analogs. The oligomers of the invention are capable of (i) forming triplexes with various target sequences such as virus or oncogene sequences by coupling into the major groove of a target DNA duplex at physiological pH or (ii) forming duplexes by binding to single-stranded DNA or to RNA encoded by target genes. The oligomers of the invention can be constructed to have any desired sequence, provided the sequence normally includes one or more bases that is replaced with the analogs of the invention. Compositions of the invention can be used for diagnostic purposes in order to detect viruses or disease conditions.

Abstract: Abundant interspersed repetitive DNA sequences of the form (dC-dA).sub.n.(dG-dT).sub.n have been shown to exhibit length polymorphisms. The polymorphisms can be used to identify individuals as in paternity and forensic testing, and can be used to map genes which are involved in genetic diseases or in other economically important traits. The polynucleotide provided consists of a DNA fragment, preferably .ltoreq.300 base pairs in length containing one or more blocks of tandem dinucleotide repeats (dC-dA).sub.n.(dG-dT).sub.n wherein n.gtoreq.6 and preferably .gtoreq.10.

Abstract: The present invention discloses nucleic acid enzymes capable of cleaving nucleic acid molecules, including single-stranded DNA, in a site-specific manner under physiologic conditions, as well as compositions including same. The present invention also discloses methods of making and using the disclosed enzymes and compositions.

Abstract: A recombinant plasmid useful as a cloning vector in lactic acid bacteria and only containing DNA of lactic acid bacterial origin, the plasmid comprising (a) a DNA fragment comprising a replication region functional in lactic acid bacteria and (b) a DNA fragment comprising a marker gene selectable in lactic bacteria, the expression of which allows one-step primary selection in lactic acid bacterial cells transformed with the recombinant plasmid; a method for constructing the cloning vector plasmid; the recombinant plasmid with an inserted gene coding for a desired gene product; a method of preparing an improved food starter culture by introducing the recombinant plasmid with an inserted gene coding for a desired gene product and food starter cultures prepared by the method are provided.

Abstract: A recombinant microorganism strain having a desired metabolic property is produced by a process which utilizes a multiple chemostat system.