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R N Bastos

Publications and source records attributed to R N Bastos.

6 recordsLinked to original sources
PubMed1979

Evaluation of several enrichment procedures for the isolation of recombinant plasmid DNA.

A number of methods for the selective enrichment of recombinant plasmids were examined; these include alkaline phosphatase treatment of the restricted pBR322 vector, as well as a combination of this and S1 nuclease treatment of the ligated mixture of pBR322 and pCR1 plasmids or S. griseus DNA followed by D-cycloserine treatment to enrich for cells carrying recombinant molecules. The relative efficiencies of these methods were compared.

Alkaline Phosphatase↗
PubMed1977

Integration and regulation of mitochondrial assembly in yeast.

The interactions between the mitochondrial and nucleocytoplasmic systems required for mitochondriogenesis have been investigated at several different levels. Those involved in the formation of functional enzyme complexes have been studied using cytochrome oxidase: this multimeric (2 X 7 and 2 X 6 subunits for enzymes from yeast and beef heart respectively) has been resolved, and the mitochondrial contribution has been shown to be dispensible for catalytic function proper. Using novel mutants, with a mitochondrial mode of inheritance, a mitochondrial gene product localized in the oligomycin-sensitive ATPase has been implicated in the assembly not only of this complex, but of cytochrome oxidase as well. Interactions required for the genetic competence of the mitochondrial system have become apparent as a result of studies in the mechanism of action of the highly effective mitochondrial mutagen ethidium bromide. This agent first becomes covalently inserted into mitochondrial DNA and, after its excision, eventually results in extensive degradation of the macromolecule. The excision reaction has now been shown to be performed by a complex between the oligomycin-sensitive ATPase and a DNA-binding protein presumably involved in recognizing the damage. On the level of replication and expression of the mitochondrial genome studies using thermolabile mutants have demonstrated that these processes appear independent of the replication of nuclear DNA but not of its expression.

DNA, Mitochondrial↗
PubMed1977

Globin RNA precursor molecules: biosynthesis and process in erythroid cells.

Hybridization of labeled RNA with excess amounts of DNA complementary to globin mRNA, in conjunction with a pulse-chase technique, were used to investigate the biosynthetic pathway of globin mRNA in erythroid cells. Three species of molecules sharing common sequences with globin mRNA were detected in the nuclei of these cells, two of which are larger than the cytoplasmic globin mRNA. One species was approximately 7 times larger than globin mRNA ("27S"), and the other ("15S") was only about twice the size of cytoplasmic globin mRNA. The largest species lacked poly(A) sequences, while the others contained poly(A), After chase, the large RNA species gradually disappeared ( 1/2 = 5 min), while the cytoplasmic 10S species accumulated. From these results a model is proposed describing the biosynthetic pathway of globin RNA transcription: an early transcription product is the large molecule "27S" (approximately 5000 nucleotides long) which is then cleaved into a smaller species "15S" (approximately 1500 nucleotides). This intermediate precursor is then clipped, presumably at the 5' end, and finally converted to the exported "10S" molecule (approximately 750 nucleotides) which accumulates in the cytoplasm.

Cell Line↗
PubMed1975

Use of diazido ethidium bromide as a specific probe for mitochondrial functions.

The diazido derivative of ethidium bromide has been synthesized as a potential photoaffinity label and shown to be at least as effective as a mitochondrial mutagen as the parent compound, with a similar mode of action. Exposure of mitochondria of Saccharomyces cerevisiae to the compound, followed by ultraviolet-irradiation, which converts it to the highly reactive dinitrene, results in its specific binding to a single component which has been tentatively identified as the smallest polypeptide (subunit 9) of the membrane-bound ATPase. An analogus reaction is also obtained with the soluble, oligomycin-sensitive ATPase complex but not with the F1-ATPase itself. The reaction with the ATPase complex can also be monitored by fluorescence enhancement and by this attribute, as well as by other criteria, diazido-ethidium bromide, ethidium bromide itself, euflavine, N,N'-dicyclohexylcarbodiimide, 2,4-dinitrophenol, and 2-azido-4-nitrophenol all appear to compete for the same, lipophilic, binding site. A mitochondrial mutation (73/1) (see Flury, U., Feldman, F., and Mahler, H.R. (1974) J. Biol. Chem. 249, 6630-6637) produces a photoaffinity product with an altered electrophoretic mobility and molecular weight.

Adenosine Triphosphatases↗