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At least 973 records · Page 54Linked to original sources
PubMed1983

Genetic applications of yeast transformation with linear and gapped plasmids.

Techniques for high frequency yeast transformation have been described. A double-strand break introduced by restriction enzyme cleavage can be used to direct a plasmid to integrate into a particular chromosomal locus. Plasmids containing a double-strand gap can be used in a straightforward method for the isolation and mapping of chromosomal alleles. These techniques extend the genetic applications of yeast transformation.

Alleles↗
PubMed1986

Consequences of widespread deregulation of the c-myc gene in transgenic mice: multiple neoplasms and normal development.

We have constructed a transgenic mouse strain in which a mammary tumor virus LTR/c-myc fusion gene is anomalously expressed in a wide variety of tissues. The deregulated c-myc transgene, now glucocorticoid inducible, contributes to an increased incidence of a variety of tumors, including those of testicular, breast, lymphocytic (B cell and T cell), and mast cell origin. The deregulated gene does not, however, otherwise disturb cell proliferation, nor does it interfere with normal development in these animals. Moreover, since not all tissues that express the transgene develop neoplasms, these results begin to define the transforming spectrum of the c-myc oncogene. They also extend to several organ systems the notion that elements in addition to an activated c-myc gene are required to induce malignancy in the living organism.

Adenocarcinoma↗
PubMed1991

Cultivation of recombinant E. coli and production of fusion protein in 60-1 bubble column and airlift tower loop reactors.

E. coli K12 with multicopy plasmid (lambda PR-promoter and temperature-sensitive lambda cI 857 repressor) was cultivated in 60-l bubble column and airlift tower loop reactors. The medium composition, cell concentration, and intracellulary enzyme activity were monitored on-line during batch, fed-batch, and continuous cultivations. The specific growth rates, cell mass yield coefficients, plasmid stabilities, productivities of the amount of active fusion protein (beta-galactosidase activity), concentrations and yields of acetic acid, and volumetric oxygen transfer coefficient were evaluated for different medium compositions and cultivation conditions. The enzyme activity was also monitored during the temperature induction. The results evaluated in the 60-l bubble column and airlift tower loop reactors are compared with those evaluated in a 1-1 stirred-tank reactor.

Bacterial Proteins↗
PubMed1983

A new technique for genetic engineering of Agrobacterium Ti plasmid.

A new technique is described that allows easy introduction of foreign genetic elements into specific regions of Agrobacterium tumefaciens DNA. It uses plasmids that (1) can be introduced, but not maintained in A. tumefaciens, (2) have a region homologous to the genome of the recipient, and (3) have an appropriate marker. Selection for the marker will yield transconjugants in which the introduced plasmid has recombined with the host genome. Applications of the technique are described.

Conjugation, Genetic↗
PubMed1986

Microcell-mediated chromosome transfer from human tumor cells to human recipient cells evidenced by premature condensation of the transferred chromosomes.

Micronucleation was induced by vincristine, colcemid, and colcemid in combination with cytochalasin B in cells of a human metastatic breast carcinoma cell line (MDA MB 231). Cells treated with the latter combination were enucleated subsequently by centrifugation in the presence of cytochalasin B. The resulting "microcell" fraction was fused with mitotic human primary fibroblasts or mitotic HeLa cells using polyethyleneglycol (PEG). The success of the microcell-mediated chromosome transfer thus could be demonstrated as premature condensation in the mitotic recipient of the transferred micronuclei. This technique of cytogenetic analysis allowed a fast and simple control of the influence of different conditions on micronucleation and fusion of micronuclei with recipient cells. It could be shown that microcell-mediated chromosome transfer from human tumor cells into human normal, as well as human tumor, recipient cells is practicable if the techniques figured out by the present study are employed.

Breast Neoplasms↗
PubMed1996

Synthesis of alpha-helix-forming peptides by gene engineering methods and their characterization by circular dichroism spectra measurements.

Two kinds of peptides which were considered to form alpha-helices were designed and characterized. One was "alpha(3)-peptide' with 21 residues comprising three repeats of the seven-residue sequence Leu-Glu-Thr-Leu-Ala-Lys-Ala. This peptide appeared to be amphipathic due to a hydrophobic surface of Leu residues and a hydrophilic surface of Lys and Glu residues, thus forming a bundle structure. The other was "alpha(3)-GPRRG-alpha(3) peptide' with 47 residues in which two alpha(3)-peptides were connected by the five-residue sequence Gly-Pro-Arg-Arg-Gly. The genes encoding these peptides were fused to the adenylate kinase gene via a methionine codon. The resulting fused protein was expressed as an inclusion body, and the peptides were purified after cleavage with BrCN. The stability of the peptides in various buffers was then examined by measuring their circular dichroism spectra. The alpha(3)-peptide showed concentration-dependent stabilization of the alpha-helix. Sedimentation equilibrium ultracentrifugation indicated that it formed a bundle structure composed of four polypeptide chains, and a dimer intermediate during oligomerization was also detected by analytical gel-filtration. The stability of the alpha(3)-peptide was decreased by shifting the pH to 2 or 12, due to electrostatic repulsion of charged residues. Thus, the alpha(3)-peptide was stabilized by increasing the ionic strength, particularly in acidic or alkaline buffer, through the masking of the repulsion by high salt concentration. In buffer of neutral pH and a high salt concentration, the alpha(3)-peptide at high concentration formed visible aggregates, due possibly to the exposed hydrophobic surfaces of the alpha-helical bundles. On the other hand, alpha(3)-GPRRG-alpha(3) peptide did not show concentration-dependent reversible dissociation and association. It was shown to exist as a trimer even at low concentration, indicating very tight association of the alpha(3)-GPRRG-alpha(3) peptide. In contrast to the alpha(3)-peptide, the alpha(3)-GPRRG-alpha(3) peptide was very stable at various pH values and salt concentrations. This seemed to be due to increased hydrophobic interactions resulting from the increase in the number of seven-residue repeats from three to six, even though each group of three repeats was separated by a five-residue sequence.

Amino Acid Sequence↗