Complementation studies

Complementation with hypF gene from T. roseopersicina in a AhypF strain of R capsulatus was successful, a clear demonstration that a functionally active form of this Thiocapsa gene product is synthesised by the R capsulatus cells from the foreign template. The same experiment using a AhypF E. coli strain resulted in barely detectable complementation. We conclude that there must be strain dependent variations in the complementation capacity and that the most thoroughly studied bacterium, E. coli, may not be the best choice for such complementation studies of hydrogenase assembly and biosynthesis. 

Complementation studies using mutant proteins in vitro provide compelling evidence that the active form of integrase must be at least a dimer [14, 17]. This can be inferred from the result that when certain inactive forms of integrase—generated either by truncation or point mutation—are mixed, robust activity can be reconstituted. This indicates that different monomers in a multimer are capable of providing different essential functions in the context of an active complex. 

Kohlbrecher, D. Eisermann, R. Hengstenberg, W. Staphylococcal phos-phoenolpyruvate-dependent phosphotransferase system molecular cloning and nucleotide sequence of the Staphylococcus carnosus ptsi gene and expression and complementation studies of the gene product. J. Bacteriol., 174, 2208-2214 (1992)... 

The interfacial capacitance can also provide a significant insight into the permeability of interfacial supramolecular assemblies. While information of this kind complements studies using redox-active probes in solution, it also provides information on a significantly shorter length scale, i.e. that of electrolyte ions and solvent molecules. For example, for dense, defect-free monolayers, the limiting capacitance is very much lower (5-10 pF cm-2) than that found for an unmodified interface (20-60 pF cm 2). 

This method can be used to compensate for inhibition of a biochemical pathway which results in a deficiency of an essential metabolic product. Detailed variations of the method are provided by Dayan et al.7 and Amagasa et al.1 The inhibitor concentration should be no higher than that required for strong herbicidal effect. Metabolite concentrations should be below that which is phytotoxic. For example, certain amino acids such at methionine, are growth inhibitors at relatively low concentrations. So, in preliminary work, dose-response studies should be done with amino acids to find the maximum concentrations that do not inhibit growth. Then, seeds of test plants should be imbibed in solutions of the phytotoxin with and without metabolite solutions. Amino acids, tricarboxylic acid cycle intermediates, vitamins, nucleotides, and reducing agents have all been used in complementation studies to elucidate modes of action of a variety of phytotoxins. Examples of each of these is provided by Dayan et al.7... 

One must be careful not to overinterpret the results of complementation studies. Sometimes reversal does not occur, even though the metabolites used are those that are depleted by the phytotoxin. This can be for many reasons, such as deregulation of the pathway by inhibition or accumulation of toxic intermediates. Reversal can also be due to direct interactions of the inhibitor with the reversing compounds.69119... 

La Vallie ER, Stahl ML, Cloning of the flagellin gene from Bacillus subtilis and complementation studies of an in vhro-derived deletion mutation, J. Bacteriol., 171 3085-3094, 1989. 

It is widely believed that I-cell disease (ML-II) and pseudo-Hurler polydystrophy (ML-III) are variants of the same disorder because fibroblasts from both these disorders are deficient in phosphotransferase activity. At the molecular level, the primary and most significant difference is that in I-cell disease the deficiency is essentially total, whereas in pseudo-Hurler polydystrophy there appears to be significant residual activity (approximately 10% of control values). The molecular basis for these two diseases may, however, be distinct, as evidenced by genetic complementation studies demonstrating complementation of ML-II by some ML-III fibroblasts (Mueller et al., 1983). Although two clinical presentations of I-cell disease have been described (i.e., the neonate and 6- to 12-month-old patient), correlation of the degree of deficiency of phosphotransferase activity with clinical severity in the two I-cell disease presentations remains controversial. 

Complementing studies of planar carbon noted in Sections 1.17.2.2 and 1.17.3.2, recent calculations have suggested molecules that should contain planar silicon. Several examples were built from Si-C-B rings <2004JA16227>. 

Once a mutant with a defined knock-out phenotype is isolated, expression of the cloned gene within the mutant is desirable in order to restore the wild-type phenotype. This is in fact one of the major motivations within the Dictyostelium research community to develop gene expression (shuttle) vectors. Such vectors, which were primarily optimized for expression of homologous genes for complementation studies, can also serve as optimized vectors for the expression of heterologous genes (see below). 

Bring to a completion the tlal mutant complementation studies Complete the biochemical analyses of the complemented strain by providing a full molecular and biochemical characterization of its properties. 

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