Showing papers in "Molecular Genetics and Genomics in 1969"

The caryological analysis of plants regenerated from tumorous and other callus cultures of tobacco

Abstract: 1. Large numbers of tumorous (auxin-autotrophic) and non-tumorous (auxin-heterotrophic) tobacco callus cultures were transferred from their culture media in darkness to media with high kinetin content in light for regeneration. There were two lines of tumorous cultures, Tc and Ta; they were started in 1946 from the tobacco variety “White Burley”. Tc had been transformed by infection with Agrobacterium tumefaciens, Ta spontaneously by “habituation”. Non-tumorous cultures (Tn1 and Tn2) were started in 1961 from an unknown variety of tobacco. All the four types of cultures we received from the laboratory of Prof. R. Gautheret in Paris, in 1965. 2. Regeneration from tumorous cultures to fully developed plants was largely successful. These plants, however, were always sterile and none represents a genuine specimen of “White Burley”. Regeneration from non-tumorous, so-called “normal”, cultures was on the contrary very poor. The number of plants regenerated from Tn2 was very small. The plants were more or less uniform, growing well only as scions on other tobacco plants. Only a few “teratomata” from Tn1 developed and, in one case only, a plantlet with roots, shoots and leaves. In grafting experiments with Tn1 teratoma as scion on “Xanthi” tobacco stock, all regenerating plants originated from material with the gene N, probably from the “Xanthi” stock. 3. A comparison of the chromosome numbers of the callus cultures with those of the plants regenerated from them shows that selection plays an important part in the process of regeneration. Plants regenerated from tumorous cultures had extreme aneuploid chromosome numbers and this fact supports the assumption that abnormal chromosome numbers of tumorous cells (in the original callus cultures) are not the cause of tumor growth. On the other hand, the high aneuploidy of the regenerated plants suffices to explain their abnormal morphology and sterility. These abnormal features should not be regarded as a consequence of the tumor condition of the original cultures. The poorly regenerating non-tumorous Tn1 and Tn2 plants, which were always abnormal and aneuploid, underline this viewpoint as do our own observations (Melchers, 1965). 4. TMV in the callus cultures (Tc and Tn2) does not influence the chromosome numbers or the ability to regenerate normal plants. In Tc nearly all regenerated plants are free of TMV, whereas all Tn2-regenerates contain TMV. This interesting difference remains unexplained. 5. “Secondary” callus cultures started from regenerated plants of Tc and Ta behave as control material of “White Burley”: on a medium with auxin, this material grows as callus, without auxin it regenerates to plantlets. 6. The experiments of Braun (1959) and his arguments against the interpretation of “recovery” (from Agrobacterium transformed cells to completely normal plants) excluding the possibility of a selection process, are discussed. His work is compared with our own and that of Lutz (1966) who described the isolation of two single cell clones, one with high, the other with low tumorous activity, from habituated material.

Cellular degeneration in the production of some mutant phenotypes in Drosophila melanogaster.

Abstract: A number of mutants of Drosophila melanogaster are characterized by the absence of structures present in the wild type. Imaginal discs from the wing mutants vestigial, apterous-Xasta, Beadex and cut and from the eye mutants Bar, eyeless and lozenge were examined by light and electron microscopy. In all these mutants, with the exception of lozenge, clear evidence of degeneration was found. The onset and duration of degeneration and the number and distribution of dying cells were specific characteristics of each mutant. In most cases the degenerate areas of the disc could be correlated with the missing parts of the adult wing or eye. In contrast, in wild type wing and eye discs and in wing discs from the mutant miniature, which has a wing reduced in size but fully formed, extensive cell death was not observed. The ultrastructural features of the degenerating areas weresimilar in all the mutants studied. Conspicuous aspects of the cytolytic process included condensation and fragmentation of the dying cells followed by phagocytosis of the cell fragments by neighboring disc cells. The results indicate that localized cell death during development is a widespread occurrence among Drosophila mutants which exhibit structural deficiencies.

A map of four genes specifying enzymes involved in catabolism of nucleosides and deoxynucleosides in Escherichia coli.

Abstract: Four genes specifying the enzymes thymidine phosphorylase, purine nucleoside phosphorylase, deoxyribomutase and deoxyriboaldolase were mapped by transduction with phage P1. All pairs show greater than 90 per cent co-transduction. The gene order was found to be dra-tpp-drm-pup, and the gene cluster was shown to lie between the hsp and ser B loci on the chromosome map of Escherichia coli.

Bichemical genetics of bacterial sporulation

Abstract: Pleitropic interactions among genes controlling the formation of bacterial spores and of sporulation-associated products are studied. In order to obtain sporulation mutants, spores have been germinated in the presence of chloramphenicol and then treated with nitrosoguanidine. In the most favorable conditions 25% of sporulation mutants have been found among the 40% surviving bacteria. This number is at least four times higher than the number of auxotrophic mutants, therefore a rough estimate of the number of genes involved in sporulation is 800. Rapid plate-tests have been developed for the oxidation of terrazolium salts, the formation of various proteolytic enzymes and the production of antibiotics. Although the exact biochemical nature of the products is not yet known, the results suggest that distinct factors, probably various enzymes (including several proteases) are detected by these tests. All of them are associated with spore formation and absent from a large number of sporulation mutants. Using these tests, the phenotypes of 500 randomly selected sporulation mutants were determined. No important differences were found between asporogenous and oligosporogenous mutants. The number of mutants deficient for several sporulation-associated characters is large, pleiotropic interactions following a defined pattern are observed. Statistical analysis indicates the existence of a unidirectional pleiotropic system. All the results agree with the hypothesis of sequential gene activation. Consequently, the sporulation-associated characters can be ordered into a linear sequence, presumably reflecting the consecutive steps in spore formation. The order obtained is the following: gelatinase, proteases acting on casein and on denatured albumin, oxidation of tetrazolium No 7, digestion of protamine, production of antibiotics (against a Staphylococcus and a Bacillus), hydrolysis of hemoglobin, oxidation of tetrazolium No 2, digestion of native albumin, synthesis of elastase. Another category of mutants, blocked in a late step of sporulation and apparently derepressed for the formation of elastase, is also described. In conclusion, arguments are put forward in favor of sequential gene activation. Sporulation genes, related by unidirectional pleiotropic interactions, form a sporulon. Generalization of this concept to other differentiating systems (a differon), its predictions and possible experimental confirmation are considered.

The influence of the reading context upon the suppression of nonsense codons.

Abstract: We have examined the response of phage T4 nonsense mutations located at various sites within the same cistron to different suppression agents. A wide range of suppression efficiency is found for both ochre (UAA) and amber (UAG) mutations under conditions where suppression provides a measurement of the amount of chain propagation past the mutated site. We have established a relationship between our measurement-the size of the phage yield-and the amount of rIIB product present in the infection. Our data suggest that the 1000-fold range of variations in yields observed in the rIIB cistron corresponds to a 30-fold range of variation in the level of rIIB product, i.e. in the relative frequency of chain propagation past the various nonsense codons included in our test. From the parallelism of response of any particular mutant to very different suppression mechanisms we conclude that the efficiency of suppression is site specific, that is to say, that the main factor determining the frequency of chain propagation at a nonsense codon by any type of suppression mechanism is the nucleotide sequence adjacent to the nonsense codon (reading context). We propose that the recognition of a natural termination signal involves a sequence longer than a nonsense codon and that nonsense codons outside of their natural environment induce variable termination rates which are reflected in the suppression potential.

Studies of the genetic mechanism of radiation-induced mitotic segregation in yeast

Abstract: Sectoring was induced with x-rays or ultraviolet in a diploid yeast strain heterozygous for seven genes located on one chromosome arm. The frequencies of sectoring of different genes were approximately linearly related to their distance from the centromere. If two or more adjacent genes sectored, the event could be explained by mitotic crossing over. Sectoring of single genes, however, was mostly nonreciprocal and resembled a conversion-type event. Approximately 80% of the sectored colonies could be explained single mitotic crossovers in one of the intergenic regions.

Biochemical and genetic studies with nitrate reductase C-gene mutants of Escherichia coli.

Abstract: Twenty-eight narC (chlC) mutants of Escherichia coli were isolated and characterised by their resistance to chlorate, inability to use nitrate as terminal electron acceptor and positive gas reaction. The extent of gas production by the majority of mutants was almost normal but quantitative differences ranging from 40 to 100% of wild-type activity were found. Biochemical studies showed that all the mutants lacked nitrate reductase, decreasing gas production was correlated with a simultaneous decrease in formate dehydrogenase activity and the lowest gas production was due to deficiencies in formate dehydrogenase and hydrogenase. The position of narC relative to other loci was determined as: purB ... hemA ... narC ... supIII,C ... galU ... attΦ80 ... tonB ... trp ... cysB by transduction analysis, and the mutant sites of 6 strains representing the complete range of gas reactions were clustered at this position. It is suggested that narC is the structural gene for nitrate reductase and the variations in phenotype may be due to polarity effects on neighbouring genes specifying components of the formate hydrogenlyase system. Transduction of narC by Φ80 could not be detected but an effect of galU− on phage P1kc susceptibility was demonstrated.

Insertion of phage Mu. 1 within prophage λ: A new approach for studying the control of the late functions in bacteriophage λ

Abstract: Insertion of phage Mu. 1 within prophage λ results in defectiveness of the λ-lysogen. When Mu is located in the genetic segment involved in the late functions, the expression of all the genes distal to the insertion is severely depressed. The results indicate that the main promotor for transcription of the late functions is located in the Q-R segment.

Two insertions in the galactose operon having different sizes but homologous DNA sequences.

Abstract: DNA from λdg phages carrying the strong-polar insertions N 102 and N 116 was transcribed into RNA in vitro. The RNA was exhaustively hybridized to λdg-DNA carrying the wildtype galactose operon. The remaining RNA contained a fraction which specifically hybridized to DNA carrying the insertion. The insertion-specific RNA hybridizes equally well to both insertions, regardless from which insetion the RNA was transcribed. The amount of insertion-specific RNA transcribed from the larger insertion N 102 is 2–3 times larger than the amount transcribed from insertion N 116.

A bacteriocinogenic factor of Enterobacter cloacae.

Abstract: Enterobacter cloacae strain DF13 produces a bacteriocin which is able to kill other strains of Enterobacter and Klebsiella. This property can be transmitted to Enterobacter cloacae strain O 2 (up to 90% of the acceptor population became bacteriocinogenic), to E. coli K12F- and E. coli K 12 Hfr. Transfer of chromosome material was never observed, suggesting that the production of the bacteriocin is determined by a plasmid. However all attempts to eliminate this plasmid failed. The plasmid F′ trp cys Col B Col V could be transferred from E. coli into Ent. cloacae DF13 and subsequently it could be eliminated by acridine orange treatment. Ent. cloacae DF13 harbours in addition two independently transferable R-factors, one determining resistance against streptomycin and sulfanilamide and the other resistance against penicillin. Most but not all Ent. cloacae O2 recombinants which have received only the bacteriocinogenic factor upon conjugation with Ent. cloacae DF 13, can transfer this property to Ent. cloacae O2 but not to E. coli. E. coli F- recombinants, which have received only the bacteriocinogenic factor cannot transfer this property. The results suggest that the bacteriocinogenic factor cannot mediate its own transfer, but can be transferred when another transmissible plasmid is present. This may be the R(str sul) factor, the F-factor in E. coli Hfr or a transfer factor (Δ) in Ent. cloacae O2. Closed circular DNA molecules were selectively isolated from these strains and investigated by electron microscopy and velocity sedimentation. Ent. cloacae DF13 harbours small closed circular DNA molecules ranging from 0.5 to 3.2 μ in contour length, 98% of which corresponds to a size class of 0.7±0.1 μ. Ent. cloacae O2 also harbours closed circular DNA ranging from 0.8 to 3.0 μ in contour length, with major size classes of 0.9 μ and 1.4 μ respectively. Circular DNA of a contour length of 3.0±0.2 μ (S20,w=26 S) corresponding to a molecular weight of 6.0×106 daltons was transferred to E. coli and Ent. cloacae O 2 concomitantly with the ability to produce the bacteriocin. A significant difference was observed in the number of copies of the plasmid between Ent. cloacae and E. coli.

The influence of heterochromatin, inversion-heterozygosity and somatic pairing on x-ray induced mitotic recombination in Drosophila melanogaster.

Abstract: Mitotic recombination has been induced with X-rays in Drosophila melanogaster larvae and assayed later as twin mosaic spots in the adult eyes. When the X-chromosomes are marked with zeste and white and the third chromosomes with roughoid and sepia, the frequency of twin spots was about 20 times higher for the X-chromosome than for the third chromosome. The greater amount of heterochromatin in the X-chromosome was considered responsible for the difference.

Biochemical and genetic classification of riboflavine deficient mutants of Saccharomyces cerevisiae.

Abstract: Twenty six riboflavine deficient mutants of Saccharomyces cerevisiae were isolated. They can be divided by biochemical methods into four classes, accumulating (i) no specific product (S), (ii) 2,5-diamino-6-hydroxy-4-ribitylaminopyrimidine (AP), (iii) 5-amino-2,6-dihydroxy-4-ribitylaminopyrimidine (HP), (iv) 6,7-dimethyl-8-ribityllumazine (LU).

Mise en évidence de plusieurs loci indépendants impliqués dans la synthèse de l'iso-2-cytochrome c chez la levure

Abstract: This study concerns the chromosomal genes controlling the synthesis of cytochrome c in yeast. In the wild type there are two molecular species of cytochrome c : iso-1 (major from) and iso-2 (minor form) which differ in many positions of their amino-acid sequence. A mutation, CY1→cy1-1, in the structural gene for iso-1, leads to iso-1 deficiency, while retaining a normal albeit small amount of iso-2-cytochrome c. The cyI-1 mutant does not grow on DL-lactate as sole carbon source, while the wild type does. This property was used for selecting cytochrome c rich revertants (CYT) from cytochrome c deficient strains cy1-1; ca 200 revertants were isolated after extensive nitrous acid mutagenesis from a haploid cy1-1 strain or from a diploid cy1-1/cy1-1 strain and ca 30 of them were analyzed genetically and biochemically. The cytochrome c of seven (CYT) revertants was extracted and characterized; none of them contained iso-1-cytochrome c, but all contained large amount of iso-2-cytochrome csufficient to compensate for the deficiency. It was concluded that none of the revertants resulted from back mutation of cy1-1 and that the cy1-1 mutation is a deletion or some other irreversible aberration. These conclusions were corroborated by genetic analysis. It was shown that every reversion is due to a chromosomal mutation segregating as a single gene. Five unlinked gene loci, CY2A, CY2B, CY2C, CY2D, CY2E, were uncovered in this way. None of them were linked to the CY1 locus. Revertants selected in the diploid strain were dominant or semi-dominant while those selected in the haploid strain were recessive. To the first class belong alleles at loci CY2A, CY2B, CY2C, while to the latter belong alleles at loci CY2D and CY2E. Five unlinked loci are implicated in iso-2-cytochrome c synthesis. Mutations selected at these loci act as suppressors of cytochrome c deficiency caused by a deletion of the CY1 locus. In fact the muations do not restore the synthesis of the deficient protein (iso-1-cytochrome c), but increase the synthesis of an another protein, structurally alike (iso-2-cytochrome c), and having very similar if not identical physiological activity. We propose the term of compensator genes to define this type of mutations. We discuss some possible mechanisms to explain the rarity of compensator mutations and the hypothesis that the locus CY2A could correspond not only to the regulatory gene for iso-2-cytochrome c but also to the structural one.

Effects of two TMV strains on the synthesis and stability of chloroplast ribosomal RNA in tobacco leaves.

Abstract: Multiplication of TMV-strains vulgare (light-green/dark-green mosaic symptoms) and flavum (severe yellow/green mosaic) had different effects on the ribosomal RNA of tobacco leaf chloroplasts. Vulgare inhibited chloroplast ribosomal RNA synthesis while having no effect on cytoplasmic ribosomal RNA synthesis (Fig. 2). Flavum inhibited chloroplast ribosomal RNA synthesis more severely than vulgare, and caused an earlier degradation of chloroplast ribosomal RNA than in control or vulgare-infected leaves (Fig. 1). Flavum also inhibited cytoplasmic ribosomal RNA synthesis. A connection between these differing effects on chloroplast ribosomal RNA metabolism and severity of visible symptoms is suggested, and discussed in relation to a possible influence on symptoms of denatured virus coat protein.

Studies on the biosynthesis of TMV. I. A system approaching a synchronized virus synthesis in a tobacco leaf.

Abstract: SummaryIt is shown by quantitative electron-microscopical analysis that veincleared leaves from plants systemically infected with TMV contain a majority of cells in the same stage of infection. Under defined environmental conditions the leaves which will show this synchronized virus synthesis can be determined at the time of inoculation. A standardized procedure for routine use of the system is described.The host-virus system has been used to investigate the sequential development of the virus-induced cytoplasmic differentiations and inclusion bodies. The rapid phase of virus particle formation commences at veinclearing and continues for about 40 hours. The first recognizable cytoplasmic change consists of a local branching and folding of the endoplasmic reticulum. This differentiation develops into the inclusion with diffuse, flexible rods found in late stages of infection together with the virus crystals.

On the functional organization of the arg ECBH cluster of genes in Escherichia coli K-12

Abstract: Among the four seemingly adjacent loci of the argECBH cluster of E. coli K-12, the last three are shown to belong to the same unit of coordinated expression; the latter exhibits a clockwise polarity in contrast to all other known E. coli operons, except the cluster governing the synthesis of the pyruvate dehydrogenase complex.

Comparison of RNA's transcribed in vivo from mitochondrial DNA of cytoplasmic and chromosomal respiratory deficient mutants

Abstract: In some respiratory deficient cytoplasmic mutants, the buoyant density of mitochondrial DNA is changed to detectable degrees, as compared to that of wild type strain: since this density shift suggests an important modification of polynucleotide sequence in mitochondrial DNA, we examined sequence homology between mitochondrial DNA of the respiratory mutants issued from cytoplasmic or chromosomal mutations. Mitochondrial DNA, nuclear DNA and total RNA were extracted from ϱ+ cells (wild type, respiratory sufficient) and from ϱ- cells (cytoplasmic “petite colonie” mutant, respiratory deficient), and molecular hybridization experiments were carried out between them. When ϱ+ RNA × ϱ+ mitochondrial DNA, formed roughly twice as much hybrids as the heterologous cross, ϱ+ RNA × ϱ1 mitochondrial DNA. Reciprocally, when ϱ- RNA was hybridized to ϱ+ and ϱ- mitochondrial DNA, the homologous cross produced again about twice as much hybrids as the heterologous cross. These results were confirmed by dehydridization-rehybridization experiments: the RNA separated from the hybrids “ϱ+ RNA × ϱ+ mit-DNA” as well as the RNA separated from the hybrids “ϱ+ RNA × ϱ- mit-DNA” were rehybridized either with ϱ+ or ϱ- mit-DNA. A preferential hybridization of ϱ+ RNA with ϱ+ mit-DNA, and of ϱ- RNA with ϱ- mit-DNA was clearly observed. On the contrary, ϱ+ and ϱ- nuclear DNA did not distinguish ϱ+ or ϱ- RNA. The same series of experiments were carried out using a chromosomal mutation,P 7 to p7, leading to the same respiratory deficient phenotype. We found that the p7 mutation did not introduce a detectable change in mitochondrial DNA base sequence. The results support the idea that the cytoplasmic hereditary factor, ϱ, resides in mitochondrial DNA and that the ϱ- mutations studied correspond to a dispersed sequence modification covering about a half of the total mitochondrial DNA genome, leaving the other half unchanged. Alternatively, the results can be explained by a hypothesis in which mitochondrial DNA is a heterogeneous population of the molecules having more or less related sequences and the mutation leads to a selection of certain molecular species. 4 S RNA was found to contain RNA species which hybridize with mitochondrial DNA. The degree of hybridization was very different for ϱ+ and ϱ- S RNA, when they were hybridized with either ϱ+ or ϱ- mitochondrial DNA.

Infectious DNA from coliphage T1. I. Some properties of the spheroplast assay system.

Abstract: DNA isolated from coliphage T1 is infective in spheroplasts of E. coli K12/1. The efficiency of the assay amounts to approximately 10-4 plaque-forming units per DNA molecule of 32·106 daltons. A linear relationship between DNA concentration and total phage yield or infective centers, respectively, holds for native DNA. For heat-treated DNA, however, the co-operation of 1.4 molecules is required for successful infection. Beyond a “critical concentration” of about 0.1μg/ml a self-inhibiting effect of infectious T1-DNA is observed. Breakage by shearing and denaturation of the DNA-molecules destroy their infectious activity. Renaturation, however, restores infectivity to 60–90 per cent of the original activity. Heat treatment of T1-DNA in M/5 NCE buffer results in narrow-coiled, mismatched molecules with partially denatured regions. Though the efficiency of infection of such molecules is reduced by about 30 per cent, the critical concentration of T1-DNA shifts to higher values by a factor of ten, thus giving an increase in the total plaque yield of the system. The effect is explained by the transition of native into narrow-coiled molecular configuration.

Mutations conferring quantitative and qualitative increases in beta-galactosidase activity in Escherichia coli.

Abstract: Sodium lactobionate is not utilized as a carbon source byEscherichia coli because it is only poorly bound and hydrolyzed by β-galactosidase and it does not induce the formation of the enzyme. However, treatment with N-methyl-N′-nitro-N-nitrosoguanidine produced 32 independent mutants able to grow on lactobionate. Most of the mutants formed β-galactosidase constitutively, 29 of them having mutations in the regulatory gene and one possibly in the operator. In addition, the mutants possessed quantitatively—or qualitatively—altered β-galactosidase. In 28 mutants the β-galactosidase activity was 1.5 to 4.5 times that of the wild-type. The enzymes of these mutants were unaltered in thermostability and substrate binding. One enzyme that was titrated immunologically possessed a molecular activity indentical with the wild-type enzyme. These mutants appear to contain extra copies of the gene for β-galactosidase. The spontaneous mutation rate to constitutivity was 6.3x10-3 and to the formation of apparently extra genes, 9.2x10-3.

A new class of arginine-requiring mutants in Chlamydomonas reinhardi.

Abstract: Forward mutations to auxotrophy have been induced in Chlamydomonas reinhardi after treatment of wild-type cells with ethyl methanesulfonate (EMS) and plating on a selective medium supplemented with yeast extract but deprived of ammonium chloride.

Intragenic complementation, hybrid enzyme formation and dominance in diploid cells of Saccharomyces cerevisiae

Abstract: 1. Intragenic complementation was studied using isoleucine requiring mutants of the is1-structural gene for threonine dehydratase in Saccharomyces cerevisiae. Twenty three mutants were induced with 1-nitrosoimidazolidone-2, four more mutants had been induced with ultraviolet light by other authors. Among the 23 chemical induced mutants there were 5 osmotic remedial mutants which could grow without isoleucine in the presence of 1M KCl, and 3 temperature sensitive mutants which expressed an isoleucine requirement only at elevated temperature. All mutants were shown to be allelic to is1. 2. Intragenic complementation was studied in all possible combinations using isoleucine independent growth as the criterion. All osmotic remedial, temperature sensitive and 6 more nonconditional mutants participated in intragenic complementation. 3. Complementation maps were constructed from data obtained at 25 and 35° C incubation temperature omitting temperature sensitive mutants. Both maps were linear but the 35° C map comprised six complementing groups whereas only four were observed at 25° C. 4. Negative complementation was studied using the three temperature sensitive mutants. It was indicated by a lack of growth at the permissive temperature (25° C) of the diploids formed by crossing temperature sensitive mutants to nonconditional mutants. Three of the five osmotic remedial mutants and seven nonconditional, non-complementing mutants showed negative complementation. 5. Enzyme assays with crude extracts of a representative sample of isoleucine requiring mutants revealed a complete absence of threonine dehydratase activity. 6. Crude extracts of well complementing mutant x mutant heteroallelic diploids were assayed for threonine dehydratase activity. Specific activities were usually very low or there was no activity to be detected at all. In some cases with a sufficiently high activity a further characterization of the enzyme was possible. The most salient feature with some complementing diploids was that feedback inhibition by isoleucine was absent under conditions where wild type enzyme was completely inhibited. 7. Threonine dehydratase activity was also investigated in mutant x wild type heterozygotes. Up to 24% of the total activity in the crude extracts of such heterozygotes was found to be resistant to feedback inhibition. This resistant enzyme fraction differed from wild type by an increased Michaelis constant and altered pH dependence. 8. Intragenic complementation is considered to result from the formation of hybrid enzymes composed of the two different types of subunit polypeptide chains coded for by the two different alleles present in the diploid nucleus. Such hybrid enzymes have properties different from the corresponding two homogeneous aggregates as shown by feedback resistant threonine dehydratase activity in heterozygous cells. 9. True dominance of an active over an inactive allele is considered to be a special case restricted to genes coding for a monomeric enzyme or to mutant alleles which code for no or only a drastically altered polypeptide chain. The normal situation is considered to be an intermediate state with an appreciable amount of hybrid enzyme formation. Hybrid enzymes can cause the appearance of new properties like feedback resistant threonine dehydratase activity. 10. More general problems like heterosis and the role of mutagenesis in diploid organisms are discussed in the light of intragenic complementation and hybrid enzyme formation.

Induction of DNA breakdown and death in Escherichia coli by phleomycin. Its association with dark-repair processes.

Abstract: Phleomycin, at concentrations above 1 μg/ml, induced breakdown of DNA and death in E. coli. Exponentially growing cultures were about 10 times more sensitive to phleomycin than were stationary cultures, and the effect was somewhat dependent on the medium.

Polarised complementation at the pyrimidine-3 locus of Neurospora.

Abstract: A re-evaluation of complementation at the pyrimidine-3 locus ofNeurospora suggests the existence of polarity. This apparent polarity is supported by polar complementation of the majority of a number of mutants induced with the acridine ICR-170, and confirmed by the behavior of an allele which is shown by reversion studies to be a frameshift mutant.

Genetic analysis of a mutant of Bacillus subtilis producingltraviolet-sensitive spores.

Abstract: A mutant ofBacillus subtilis, uvssp-42-1, producing UV-sensitive spores was studied genetically. By treatment of the cells with DNA prepared from auvr strain two types,uvs-42 (Hcr−) andssp-1 (Hcr+), of transformants producing UV-resistant spores were obtained. Only strains having both types of mutations together produced UV-sensitive spores.

Studies on the functions of the RNA polymerase components by means of mutations.

Abstract: It had been shown earlier, that RNA polymerase 13 S particles contain the large components with a molecular weight of about 3–105 and small subunits with a molecular weight of 4·104-1·105. These polymerase components easily dissociate and reassociate with restoration of the enzyme activity.

Classification and intragenic position of mutations in the beta-galactosidase gene of Escherichia coli.

Abstract: Following treatment with N-methyl-N′-nitro-N-nitrosoguanidine, 1,257 mutants of Escherichia coli K12 were isolated on lactose-tetrazolium medium. Of these mutants, 345 were lactose-negative and lacked appreciable β-galactosidase activity. About half of these enzyme-deficient mutants had lost the whole lactose operon; the remainder (174) were point mutations within the β-galactosidase gene. With the exception of 42 which could not be classified, the mutations were identified either as chain-terminating (UAG 57, UAA 6, UGA 60) or missense (9). There were no mutations of the reading frame and no short deletions. The unclassifiable mutants do not form crossreacting protein and are probably a type of chain-terminating mutant.

Studies with α-ketoglutarate dehydrogenase mutants of Escherichia coli

Abstract: Two classes of mutant lacking α-ketoglutarate dehydrogenase complex activity were detected by biochemical analysis of strains of Escherichia coli requiring succinate for aerobic growth on glucose minimal medium. One class, designated sucA, lacked the α-ketoglutarate decarboxylase component (E1) whereas the other class, sucB, lacked the dihydrolipoyl transsuccinylase component (E2). Studies with mixed cell-free extracts showed that the overall dehydrogenase activity could be reconstituted from several pairs of sucA plus sucB mutants but not from mixtures of mutants of the same class. Transduction analysis with phage P1 indicated close linkage between the two genes and their frequencies of cotransduction with gal were similar. The order of the two genes was also established as sucA (E1)-sucB(E2)...gal by reciprocal three-point crosses with several pairs of mutants.

Dominance and recessiveness at the protein level in mutant x wildtype crosses in Saccharomyces cerevisiae

Abstract: The is 1-locus of the yeast Saccharomyces cerevisiae is the structural gene for threonine dehydratase. is 1-mutants require isoleucine for growth and do not have active threonine dehydratase. Interallelic complementation is frequent among is 1-mutants. This is indicative for an aggregate or multimeric structure of yeast threonine dehydratase. Complementing and non-complementing mutants were crossed to wildtype. Properties of threonine dehydratase were assayed in crude extracts of the resulting heterozygotes. Specific activities varied considerably between full wildtype activity and a level about 10% of that. The apparent Michaelis constants were increased in many heterozygotes. This effect was probably due to the aggregation of both mutant and wildtype subunits to form a hybrid threonine dehydratase with reduced substrate affinity in addition to pure wildtype enzyme. This notion is supported by the observation in one heterozygote of two enzyme fractions with increased Michaelis constants in addition to a wildtype-like fraction. The possible formation of hybrid enzymes with normal, reduced or no activity is considered to blur gene dosage relations. A given pair of alleles in a heterozygous cell can generate a new type of enzyme with properties not encountered in the corresponding two homozygous cells. This situation is not accounted for by the classical concepts of dominant-recessive or intermediate behaviour, because the difference between the heterozygotes and the homozygotes is not necessarily only quantitativ but also qualitative.

Integration defective mutants of bacteriophage P2

Abstract: Mutants of phage P2 unable by themselves to be integrated as prophages have been isolated. These mutants (int) are complemented by the wild type allele and may then yield stable lysogenic strains carrying an int prophage at location I in Escherichia coli C. These lysogens produce either no phage or little phage, depending on the int mutant used. All int mutants isolated appear to belong to a single complementation group.

Genetic studies of the ribosomal proteins in Escherichia. coli. II. Altered 30s ribosomal protein component specific to spectinomycin resistant mutants.

Abstract: SummarySpectinomycin resistant (spcr) mutants were obtained by treating the cells of E. coli K12, W3637 with nitrosoguanidine. The compositions of ribosomal proteins were analyzed for six out of eleven such spcr-mutants with chromatography on a carboxymethyl cellulose (CMC) column. The 30s ribosomal subunit from all of the spcr-mutants was found to contain the altered 30-4 protein component, while no difference was detected in 50s ribosomal proteins between spcr and spcs bacteria.