Abstract: A construct which includes a cap independent 5' noncoding region of viral or cellular origin and a nucleotide sequence of interest, which is located downstream of the noncoding region. A method of producing a protein or a polypeptide of interest by introducing the construct, including a nucleotide sequence encoding the protein or the polypeptide of interest, into mammalian cells is also described. In one embodiment, the construct comprises all or a portion of the poliovirus cap independent 5' noncoding region and a nucleotide sequence encoding a protein or a polypeptide of interest.

Abstract: A novel method that allows introduction of unidirectional deletions into cloned DNA is described. This method is based on the use of a mixture of oligodeoxynucleotide primers that have fixed 5' (or 3') ends defining the end point of the deletion and variable 3' (or 5') ends composed of mixtures of all four nucleotides at six positions. The 5' ends of the oligodeoxynucleotides are hybridized to a fixed location of the M13K11RX templates and the 3' ends are hybridized randomly to the DNA to be analyzed. Such oligodeoxynucleotide primers when extended with DNA polymerase can direct deletions of intervening parts of the single-stranded DNA that by design contains multiple Eco K sites; the deletion products are selected on a host strain with the Eco K restriction system (e.g., using JM101 cells). This method is an efficient way of generating a nested set of deletion mutants useful for dideoxy-sequencing. It can also be used for creating a set of deletion mutants with a particular codon at the 5' or 3' end point.

Abstract: The invention relates to nucleic acid segments useful in the construction of expression vectors for expression of heterologous polypeptides directed to particular areas of the host cell. Selected constructs direct production of polypeptides to the outer membrane surface of the cell. Other constructs direct expression of heterologous polypeptides to the inner membrane/periplasm of the host cell. Transformed host cells are potentially useful for the production of vaccines or immunogens elicited in response to antigens expressed on the outer membranes of the host cells.

Abstract: Non-tumorigenic, human bronchial epithelial cell lines are provided wherein the cell lines are capable of expressing cytochrome P450 genes which have been inserted into the cell lines. Also provided are methods and kits for identifying potential mutagens, cytotoxins, carcinogens, and chemotherapeutic agents utilizing these cell lines.

Abstract: A process for the production of an E. coli strain producing high yields of 7.beta.-(4-carboxybutanamido) cephalosporin acylase, comprises: (a) digesting the DNA of a microorganism whose DNA includes the sequence encoding 7.beta.-(4-carboxybutanamido) cephalosporin acylase, and forming a plasmidic library; (b) transforming, with the sequences in the plasmidic library, an auxotrophic E. coli host; (c) selecting for transformed E. coli hosts containing the acylase sequence by growth on a suitable medium; (d) isolating the vector containing the acylase sequence, digesting the vector, and ligating the DNA sequences obtained into an E. coli vector under control of an E. coli promoter; (e) repeating selection procedure of step (c) ; (f) using the vectors from the selected E. coli hosts to transform an E. coli host lacking substantial .beta.-lactamase activity. 7-Aminocephalosporanic acid and its derivatives can be prepared by reaction of substrates like 7.beta.-(4-carboxybutanamido) cephalosporanic acid with 7.beta.

Abstract: The present invention provides transfer-deficient pAgK84 plasmids, non-pathogenic strains of Agrobacterium radiobacter K84 carrying such plasmids, and a method useful for control of crown gall disease mediated by A. tumefaciens. The plasmids include genes encoding the synthesis of the antibiotic agrocin 84 and are modified in the transfer region of the plasmid by sufficient deletion to inhibit transfer of the plasmid. The plasmids are used to develop non-pathogenic strains of A. radiobacter K84 that stably produce agrocin 84 and inhibit the development of crown gall disease.

Abstract: RNA enzymes or ribozymes can act as endoribonucleases, catalyzing the cleavage of RNA molecules with a sequence specificity of cleavage greater than that of known ribonucleases and approaching that of the DNA restriction endonucleases, thus serving as RNA sequence specific endoribonucleases. An example is a shortened form of the self-splicing ribosomal RNA intervening sequence of Tetrahymena (L-19 IVS RNA). Site-specific mutagenesis of the enzyme active site of the L-19 IVS RNA alters the substrate sequence specificity in a predictable manner, allowing a set of sequence-specific endoribonucleases to be synthesized. Varying conditions allow the ribozyme to act as a polymerase (nucleotidyltransferase), a dephosphorylase (acid phosphatase or phosphotransferase) or a sequence-specific endoribonuclease.

Abstract: EHS murine tumor cells express basement membrane proteins such as laminin, entactin, collagen and proteoglycan. An immortal mammalian cell line designated BAM having phenotypic characteristics of EHS murine tumor cells may be prepared by culture of EHS cells in nutrient culture medium supplemented with low concentration of mammalian serum. BAM cells express basement membrane proteins in vitro.

Abstract: Simplified methods to produce recombinant adeno-associated virus (rAAV) vectors are described. The methods involve the use of chimeric plasmids which incorporate the Epstein Barr nuclear antigen (EBNA) gene, the latent origin of replication of Epstein Barr virus (oriP), and a rAAV genome. The chimeric plasmids themselves are also a part of the present invention. These plasmids are maintained as multicopy extra-chromosomal elements in cells, such as human 293 cells. Permanent cell lines carrying these EBV/AAV plasmids are induced to produce large amounts of rAAV upon addition of wild-type, adeno-associated virus helper functions. Vectors produced in this manner are capable of transducing exogenous genes into other human cell lines and exhibit the attributes of vital elements produced by conventional methods.

Abstract: Insect control agents comprising genes encoding proteins affecting the growth, development or behavior of an insect are provided. The gene is either activated to prevent insect molting and pupation or is inactivated to reduce the feeding behavior, inhibit growth and result in the earlier death of the insect host. Such an insect control agent is exemplified by a baculovirus in which the gene encoding ecdysteriod glucosyltransferase has been inactivated. Additionally, such baculoviruses may be further modified to express a protein which affects ecdysis. Methods for producing the insect control agent and methods of controlling insects by exposing them to the insect control agent are also included.

Abstract: A process for cloning the chitinase gene of Vibrio parahemolyticus is provided, comprising the steps of cleaving the Vibrio parahemolyticus DNA with Sau3A, PSTI or other restriction enzyme, mixing the cleaved DNA fragments in the presence of pUC18 and T4 ligase to produce a composite plasmid, and inserting the composite plasmid in a DH5a strain of E. Coli.

Abstract: Isolated DNA or RNA molecules capable of hybridizing under stringent conditions to the myoD regulatory region, its proximal promoter and distal enhancer regulatory regions, and regulatory elements within the proximal and distal regions for binding basic helix-loop-helix proteins, MyoD, and proteins binding at SP1, AP1, CAAT, M-CAT, CArG, and MEF sites in DNA. DNA or RNA expression vectors for introducing a gene into a cell under the regulatory control of the myoD regulatory region. Transduced or transfected pre-muscle cells, myocytes, or myoblasts. Methods of inducing a muscle phenotype in a non-muscle cell, positively selecting for a cells expressing MyoD, and negatively selecting for cells expressing MyoD.

Abstract: The present disclosure provides the complete primary amino acid, and underlying DNA, sequence for the 47-kilodalton surface immunogen of Treponema pallidum, subsp. pallidum. The sequence was obtained by using a combined strategy of DNA sequencing of the cloned gene as well as confirmatory N-terminal amino acid sequencing of the native antigen. An open reading frame corresponding to the 47-kDa antigen was comprised of 367 amino acid codohs, which gave rise to a calculated molecular weight for the corresponding antigen of about 40,701. Also disclosed are methods for preparing variant and mutant molecules having biologically similar attributes, as well as methods for preparing particular antigenic/immunogenic subportions of the 47-kDa protein. In particular aspects, antigenic/immunogenic subportions are identified by hydrophilicity analysis of the protein sequence.

Abstract: Yeast promoters of glycolytic enzymes are modified by isolating a fragment encompassing the RNA polymerase binding site and joining to the 5' end of this fragment a DNA sequence providing for enhanced inducible or constitutive transcription of a structural gene. Constructs are prepared for efficient expression of foreign genes in yeast.Yeast strains 2150-2-3(pC1/1GAPSOD) and AB110(pC1/1GAPATi9), producing human .alpha..sub.1 -antitrypsin and superoxide dismutase, were deposited at the A.T.C.C. on May 9, 1984 and given Accession Nos. 20708 and 20709, respectively; and 2150-2-3(GAP5), 2150-2-3(Pyk5) and 2150-2-3(PHO5GAP1), expressing Hepatitis B surface antigen, were deposited at the A.T.C.C. on May 9, 1984 and given Accession Nos. 20705, 20706 and 20707, respectively.

Abstract: The present invention comprises novel recombinant DNA compounds which encode the .about.40,000 dalton adenocarcinoma antigen recognized by monoclonal antibody KS 1/4. Eukaryotic and prokaryotic expression vectors have been constructed that comprise novel KSA-encoding DNA and drive expression of KSA when transformed into an appropriate host cell. The novel expression vectors can be used to produce KSA derivatives, such as non-glycosylated KSA, and to produce KSA precursors, such as nascent KSA, and to produce subfragments of KSA. The recombinant-produced KSA is useful for the diagnosis, prognosis and treatment of disease states including adenocarcinomas of the lung, prostate, breast, ovary and colon/rectum; and for the creation of novel antibodies for treatment or diagnosis of the above.

Abstract: A method for producing infectious recombinant baculoviruses in bacteria is described. A novel baculovirus shuttle vector (bacmid) was constructed that contains a low-copy-number bacterial replicon, a selectable drug resistance marker, and a preferred attachment site for a site-specific bacterial transposon, inserted into a nonessential locus of the baculovirus genome. This shuttle vector can replicate in E. coli as a plasmid and is stably inherited and structurally stable after many generations of growth. Bacmid DNA isolated from E. coli is infectious when introduced into susceptible lepidopteran insect cells. DNA segments containing a viral promoter driving expression of a foreign gene in insect cells that are flanked by the left and right ends of the site-specific transposon can transpose to the attachment site in the bacmid propagated in E. coli when transposition functions are provided in trans by a helper plasmid.

Abstract: An E. coli heterologous gene expression system comprising a multicopy expression vector comprising an F promoter from bacteriophage P2; and at least one copy of a DNA sequence comprising a delta gene from satellite P4 operably linked to a regulatable promoter on a different replicon is disclosed.

Abstract: For the detection of nucleic acids of definite sequence by hybridisation with a complementary nucleic acid probe which contains bound via a chemical bonding at least one hapten as labelling one uses, as hapten, a steroid which is bound via a bridge of at least 4 atoms length to at least one position of the nucleic acid probe which does not participate in hydrogen bridge formation and detects the hybridised probe via an in turn labelled anti-hapten antibody.

Abstract: The invention relates to an expression system which permits control of the levels of protein produced, and optionally provides production of the mature form of the protein directly. The expression systems of the invention include the desired gene downstream of a first DNA which comprises the reverse transcript of an inducible translational regulator and a second DNA which is the reverse transcript of an mRNA capable of self-stabilization. This translated sequence is in turn under the control of a transcriptional promoter which may also be inducible. In a preferred embodiment, the inducible translational regulator is the iron responsive element (IRE) region of ferritin mRNA, and the stabilizing element is also a portion of the ferritin sequence. The expression system is useful, especially, for the production of toxic proteins since protein production can be delayed until desired.

Abstract: A novel plasminogen activator which is identical in peptide sequence to naturally occurring human prourokinase except that the 155th amino acid counting from the N-terminal amino acid (serine)] is other than the 155th amino acid (proline) of naturally occurring human prourokinase, optionally with the addition of methionine at the N terminus.