Alcohol dehydrogenase isoenzymes fromNicotiana tabacum include ADH of bothN. sylvestris andN. tomentosiformis

Electrophoretic separation of seed alcohol dehydrogenase (ADH) fromNicotiana tabacum on 12% starch gels at pH 7.8 produced only one band with an apparent R_f of 0.65, which confirmed earlier reports. The same was found with pollen ADH. However, in polyacrylamide gel isoelectric focusing, seed ADH separated into three distinct bands with apparent pI of 5.33, 5.42 and 5.50. The pI 5.33 isoenzyme was found to be the essential form inN. sylvestris seeds. The analysis of charge properties ofN. tomentosiformis seed ADH showed only one isoenzyme with pI of 5.56. These results present further evidence thatN. tabacum has arisen from a cross between aN. sylvestris predecessor and an ancestral type ofN. tomentosiformis. The presence of the pI 5.42 form inN. tabacum is consistent with the reported formation of heterodimeric ADH in tobacco hybrids.     

Flow cytometric estimation of nuclear DNA amount in diploid bananas (Musa acuminata andM. balbisiana)

Cell nuclei were isolated from leaf tissues of wild banana (Musa balbisiana, M. acuminata ssp.banksii andM. acuminata ssp.errans) and of the two vegetative clones of diploid cultivar “Pisang Mas”. Relative fluorescence intensity was measured on propidium iodide-stained nuclei by flow cytometry. Nuclei isolated fromGlycine max with known nuclear genome size were used as internal standard to determine nuclear DNA content ofMusa in absolute units. The results of the study showed that the size of nuclear genome ofMusa is smaller than previously estimated. In general, it is smaller in comparison with many other angiosperms. Furthermore, it was found that nuclear DNA content ofM. balbisiana (genome BB) is significantly lower than that ofM. acuminata subspecies and cultivars (genome AA). This finding should permit estimation of genome composition in triploidMusa clones with expected hybrid composition. Flow cytometry is proposed as a useful technique with potential applications in taxonomy, breeding and biotechnology ofMusa.     

Cytological relationships of selected species ofPanicum L.

The cytological investigation of 12 taxa ofPanicum L. revealed that the vast majority of them have the basic number x=9 at different ploidy levels. The basic number x=8 was recorded only in the tetraploid speciesP. maximum with 2n=32. The diploid number 2n=18 was encountered inP. capillare, P. laevifolium, P. antidotale andP. coloratum (2) with 3B-chromosomes recorded in the latter species. The tetraploid chromosome number 2n=36 was found to exist inP. miliaceum, P. miliare, P. coloratum (1) andP. virgatum. The hexaploid number 2n=54 was recorded inP. bulbosum, P. dichotomiflorum andP. esculentum. The karyotypes of all accessions were mostly symmetrical and mainly comprised of meta- and submetacentric chromosomes with little variation in length among them within each karyotype. Investigation of chromosome association during metaphase I of meiosis revealed that the frequency of bivalents/cell was the highest among all investigated diploid, tetraploid and hexaploid accessions. Univalents were also frequently encountered in various accessions. These results may indicate that segmental alloploidy has been the major process by which polyploid species have originated.     

Crop/Weed gene flow:Cucurbita argyrosperma Huber andC. fraterna L. H. Bailey (Cucurbitaceae)

A mixed population of Cucurbita at Vado El Mow in northern Tamaulipas, Mexico showed an anomalous pattern of fruit bitterness. Some domesticated plants (C. argyrosperma andC. moschata) expressed cucurbitacin bitterness whereas some sympatric free-living plants produced non-bitter fruits. This reversal of typical cucurbitacin expression suggested gene flow between crop and weed at the site. Isozyme analysis provided little insight as to taxa involved in gene exchange, although progeny from a single free-living plant carried IDH allozymes that are associated with Mexican landraces ofC. pepo. Synthetic hybridization revealed that fertile F, hybrids are produced from crosses involvingC. fraterna as the pistillate parent andC. argyrosperma as the staminate parent. Interspecific crop/weed hybrids can produce viable progeny upon self-pollination or backcrossingto either parent, andF₂ families display normal allozyme segregation. Hybrid fertility, as indicated by pollen stainability, increases in progeny produced by backcrossingfrom theC. argyrosperma parent. Interspecific hybridfertility represents a potential for crop/weed gene flow that would be realized under natural conditions if pollen flow occurs betweenC. fraterna andC. argyrosperma in the fields of Tamaulipas. Oligolectic “squash bees” (Teponapis), efficientCucurbita pollen vectors, are present at the site. Thus, it is likely that natural interspecific crop/weed hybridization has occurred at Vado El Moro and this might at least partially explain the anomalous distribution of fruit bitterness among extant populations at the site.     

Characterization of mitochondrial DNA topoisomerase I fromNeurospora crassa

DNA topoisomeraseI isolated from the lower eukaryoteNeurospora crassa mitochondria was characterized. Molar mass of the enzyme in the native state is 120 kDa and 60–65 kDa when denatured. The pH optimum of the enzyme is 7.8 and the KCl optimum concentration is 40 mmol/L. This topoisomerase is independent of ATP and Mg²⁺. N-Ethylmaleimide, 4-chloromercuribenzoate, SDS, guanidinium chloride, polyethylene glycol, heparin and ethidium bromide inhibit its activity, while novobiocin, nalidixic acid, Triton X-100 and chloroquine do not. Polyamines, and histone, H1 stimulate the topoisomerase activity. We classify this DNA topoisomerase as typeI and eukaryotic. Conversion of the topoisomerase to a nonspecific endonuclease at increased temperature is proposed.     

Subcellular localization of ribonuclease isoenzymes in tobacco mesophyll protoplasts and their changes induced by infection of PVY

Changes in the content and subcellular localization of ribonuclease isoenzymes were determined in mesophyll protoplasts prepared fromNicotiana tabacum L. cv. Samsun from healthy and potato virus Y (PVY) infected plants. Intact chloroplasts, mitochondria and soluble cytosolic proteins were obtained after protoplast disintegration by means of differential centrifugation. The 1 000g pellet from healthy protoplasts contained 7.3 %, the 15 000g pellet 13.5 % and 15 000g supernatant 82.1 % of the total activity of ribonucleases. The 1 000g pellet from infected protoplasts contained 10.4%, the 15 000g pellet 10.0% and 15 000g supernatant 89.6 % of the total activity of ribonucleases. The activity of these enzymes in infected protoplasts was enhanced in crude homogenate to 137.0 % (P<0.001), in 1 000g pellet to 194.8 % (P<0.001), in 15 000g pellet to 101.3 % (NS), and in 15 000g supernatant to 149.4 % (P<0.001) of that in healthy noninoculated protoplasts.     

Plant radioresistance and DNA repair efficiency inChlamydomonas reinhardtii andPisum sativum

This paper compares the repair of DNA single strand breaks (ssb) induced by γ-radiation in two strains ofChlamydomonas reinhardtii (137C/+/ and UVS-I) and three lines ofPisum sativum (NN 131, 198, 140) differing in the degree of radioresistance. DNA ssb in cells exposed to γ-rays (50, 100, 200, 500 Gy) were measured by electrophoresis and alkaline unwinding method with subsequent chromatography on hydroxyapatite immediately after irradiation and after 30 min of post-irradiation incubation at 25°C. An increase of double-strand DNA (in%) was found in cells after 30 min post-irradiation incubation.C. reinhardtii strains displayed an equal level of DNA degradation and repair efficiency in the DNA single strand breaks. The radioresistant line N 198 ofP. sativum is characterized by a lower level of induced DNA ssb and higher efficiency of repair of these breaks as compared with less radioresistant lines NN 131 and 140.     

Restriction endonucleases R.EcoKI and R.EcoR124I are probably located in different environments within the bacterial cell

We describe the phenomenon of a transient state of R124I restriction deficiency after long-term storage of theE. coli[pCP1005] strain at 4°C, or after growth of the culture in synthetic M9 medium with the nonmutagenic solvent dimethyl sulfoxide. The unusual high reversion from the R⁺ ₁₂₄ to the R^? ₁₂₄ phenotype was observed only inE. coli strain transformed with the high-copy number plasmid pCP1005 carryingECoR124IhsdR, M and S genes cloned, but not with strains carrying the natural conjugative plasmid R124. The effect of both treatments on the expression ofEcoR124I phenotype in relation to the possible location of R.EcoR124I restriction endonuclease inE. coli is discussed.     

Morphology of the slashed pod trait in lentil

Reducing losses from pod shatter is a major goal of most lentil (Lens culinaris) improvement programs, however, genetic variability is limited. Recently, a slashed pod trait was suggested to have potential value for reducing losses from shattering, but little was known about the trait. In this study we determined the anatomical features which were associated with the slashed pod trait. Histological specimens from ‘Brewer’ lentil (normal pods) and from germplasm lines expressing the slashed pod trait were compared to each other and to specimens from normal and reduced pod parchment pea (Pisum sativum) lines. Reduced parchment pea pods had less sclerenchyma tissue and fewer fibers than pea pods with normal parchment, but all lentil pods examined had comparable sclerenchyma tissue with similar quantities of fiber. The slashed pod trait was not the result of reduced sclerenchyma tissue (parchment) as had been previously suggested. Apparently, the slashed pod trait resulted from the stresses which develop between fiber cells within the sclerenchyma layer of the pod wall during pod maturation and drying. The trait had little effect on quality of seeds for human utilization in the environments tested but may have an undesirable effect in other environments. Although seed loss due to shatter appeared to be decreased in pods exhibiting the trait, the uncertainty of expression due to environmental influences makes the trait an unlikely candidate for use in lentil improvement programs.     

Proline metabolic pathways in calli fromLycopersicon esculentum andL. pennellii under salt stress

The enzyme activities of the proline metabolic pathways were determined in control and satt-treated (140 M NaCl) calli derived from cotyledons of the domestic saltsensitive tomatoLycopersicon esculentum and the wild salt-tolerantL. pennellii. Glutamate, glutamine, asparagine, and aspartate levels increased in both genotypes under salt stress, while proline accumulation increased markedly only in the salt sensitive tomato. Activity of glutamine synthetase (GS) decreased in the salt-treated calli of the domestic species, whereas both NADH- and NADPH-glutamate synthase (GOGAT) activities increased; GS and NADPH-GOGAT decreased together in the salinized calli of the wild species. Decreasing ornithine levels were found due to NaCl in both tomato populations, while ornithine transaminase (OT) decreased in the wild type only. Increasing NADPH-Δ-pyrroline-5-carboxylate reductase (P5CR) and decreasing proline oxidase (Pro oxi) occurred in the salinized calli of the wild type. Conversely, Pro oxi and proline dehydrogenase (Pro dH) decreased highly in the salinized calli of the domestic population, while no significant changes in P5CR were found.     

Effects of inorganic arsenicals on DNA synthesis in unsensitized human blood lymphocytes in vitro

Effects of inorganic arsenicals on DNA synthesis in unsensitized human blood lymphocytes were biphasic: The chemicals at very low concentrations enhanced DNA synthesis, whereas higher concentrations inhibited DNA synthesis. The concentrations of arsenicals at which the maximum stimulating effect was found were 1×10^?5 M, 1×10^?6 or 2×10^?6 M, and 0.8×10^?6 or 1×10^?6 M for sodium arsenite exposure of 1 h, 3 d, and 6 d, respectively; for sodium arsenate, 1× 10^?5 M, 1×10^?5 M, and 2×10^?6 or 5×10^?6 M, respectively. Arsenicals must be present for the entire 6-d culture period to produce maximum stimulation of DNA synthesis in human lymphocytes. The longer exposure of the lymphocytes to arsenicals, the lower the concentrations of arsenicals at which the maximum stimulating effect on DNA synthesis was found. Stimulating effect of trivalent arsenic (sodium arsenite) on DNA synthesis was stronger than pentavalent arsenic (sodium arsenate), and the stronger the effect of trivalent arsenic than pentavalent, the longer exposure of the cells to the chemicals. Both sodium arsenite and sodium arsenate stimulated DNA synthesis in human lymphocytes to a lower degree than phytohemagglutinin (PHA).     

Seed germination of the Australian desert shrubEremophila (Myoporaceae)

Eremophila is an Australian genus of 212 species ranging from prostrate shrubs to small trees, the great majority of which occur in Western Australia. Recent interest in the genus’s germination strategies developed out of a need to seek rehabilitation techniques for mine-site and rangeland areas. The genus is of special interest because of its broad geographic range and prominence in vegetation associations of the arid zone, especially in Western Australia, where it often dominates or codominates over wide areas. ManyEremophila species are known to be tolerant of drought, fire, frost, grazing, and salinity, and would be very suitable for revegetation programmes; however, the genus germinates poorly, for reasons that are not fully understood. During the early 1980s attempts were made, with limited success, to rehabilitate open-cut mine-site waste dumps at Mt. Newman using broadcast seed and rooted cuttings and transplanted seedlings at the Pilbara mine venture at Paraburdoo. Tissue culture and grafting have also been attempted, but environmental management acknowledges that the broadcast seed method is the only cost-effective means of mine-site rehabilitation. Under field conditions the germination ofEremophila occurs in response to heavy rain in autumn and winter, especially with milder temperatures; however, up until now the use ofEremophila in rehabilitation practices has been limited because of the high percentage of seeds that fail to germinate. Two causes for this have been postulated. First, the environmental conditions may not be appropriate for germination. Physical dormancy resulting from the hard woody fruits may be overcome by using a scarification pre-treatment. Second, seed may still fail to germinate even though favourable conditions exist; a secondary chemical mechanism in the form of inhibitors associated with the fruit wall is hypothesised. Chemical properties of the seed, seed coat, or fruit may regulate germination until the fruit wall has been effectively worn away and the chemicals leached out. Few experimental procedures have been carried out to verify these hypotheses, and few studies have examined either fruit productivity (seeds per fruit) or seed viability. Eremophila fruit are dry with a papery exocarp or drupaceous with a fleshy or succulent mesocarp and a woody or crustaceous endocarp and contain between 2 and 12 seeds. Seed viability inE. maculata ranges from 74% to 92% in the first 3 years after fruit maturity, decreasing markedly to 8% after 13 years. Similar patterns have been recorded forE. goodwinii. Effective environmental management utilisingEremophila may be approached in three ways. First, scarify fruit to promote the uptake of water and oxygen; second, use fruit between 1 and 5 years to ensure the highest viability rating; and third, collect fruit from healthy shrubs showing no evidence of insect or fungal attack to ensure quality of fruit. Further research on the genus should include ecological studies, currently poorly understood, and edaphic and soil amelioration projects (e.g., on post mine-site tailings) in an attempt to optimise the vegetation potential ofEremophila.     

Mitochondrial DNA restriction maps ofMus booduga,Mus terricolor andMus musculus tytleri

Restriction maps of milochondrial DNA of the Indian pygmy field miceMus booduga andM. terricolor, and the house mouseM. musculus tytleri were determined with seven six-cutter restriction enzymes. The restriction map of the mitochondrial DNA of the laboratory mouseM. m. domesticus was used as a reference. Pairwise comparison was made of the mitochondrial DNAs for the presence or absence of the restriction sites, and per cent sequence divergence was calculated. The results show that the sequence divergence betweenbooduga andterricolor is 8-7% while thedomesticus-tytleri andbooduga-terricolor groups are divergent by 16-3%. The mtDNA sequence divergences we have obtained suggest that thebooduga-terricolor lineage might not have diverged before the Southeast Asiancervicolor-cookii-caroli lineage during evolution of these lineages of the subgenusMus as inferred by others earlier. On the other hand, it seems likely that these lineages evolved in parallel.     

Diversity in two candidate malaria vaccine antigens in different Indian strains ofPlasmodium falciparum

Polymorphism in two malarial antigens, merozoite surface antigen-1 (MSA-1) and ring erythrocyte surface antigen (RESA), has been characterized in four different Indian strains ofPlasmodium falciparum. The Indian strains were obtained from two malaria endemic regions of India, viz. Surat (Gujarat) and Delhi, and established in culture. Monoclonal and polyclonal antibodies raised against different domains of these antigens were used in the study. In MSA-1 a novel intragenic crossover was detected in the central conserved domain in two of the Indian strains. The repeat domain of RESA was found to be absent in the two strains ofP. falciparum isolated from Surat. These differences in immunoreactivity have been extended to the DNA level by appropriate PCR studies. MSA-1 and RESA are candidate vaccine antigens and these diversities will have an important bearing on the design of a suitable malaria vaccine.     

Hormone-like product of vascular tissue stimulating starch accumulation in pith explants of kale and endogenous cytokinins

When stem explants of kale (Brassica oleracea L. var.medullosa), containing pith parenchyma and a strip of vascular tissue, were cultured on simple sucrose medium, a hormone-like factor was transported from the vascular tissue to the adjacent pith, where it stimulated accumulation of starch. Similarly, up to a sevenfold increase of starch content in explants could be induced by cytokinins added to the culture medium. The relative stimulatory effect of several cytokinins (5×10^?6 M) and hormone-like product of vascular tissue (HPVT) in a typical experiment were: control (1.0), trans-zeatin (6.7), HPVT (6.2), N⁶-[2-isopentenyl]adenine (5.4), transzeatin riboside (5.2), N⁶-[2-isopentenyl]adenosine (5.4), kinetin (3.6), 6-benzylaminopurine (3.5), and adenine (2.1). Concentration of endogenous cytokinins was determined using ELISA (trans-zeatin, N⁶-[2-isopentenyl]adenine and their ribosides) andAmaranthus bioassay (total cytokinins). No effect of vascular tissue on the level of endogenous cytokinins in explants was found. The results support the conclusions of previous experiments that the HPVT stimulating starch accumulation is not a cytokinin.     

Occurrence of genes for P and S fimbriae and hemolysin in urinaryEscherichia coli

Escherichia coli is the common causative agent of urinary tract infections. Sixty-one strains ofE. coli isolated from children with urinary tract infections were tested by colony hybridization for the presence of genes determining P and S fimbriae and hemolysin. Of these strains, 46 possess a gene for hemolysin, 44 for P fimbriae and 28 for S fimbriae. Only 30 strains formed lytic zones around the colonies on plates with sheep erythrocytes. The results indicated that simultaneous occurrence of genes in urinaryE. coli was highest for P fimbriae and hemolysin and lower for other combinations of the tested genes.     

Superoxide dismutase level in response to paraquat and high temperature in the cyanobacteriumGloeocapsa sp

Paraquat (methylviologen) at concentrations above 0.05 mM inhibited the growth of photoautotrophic cyanobacteriumGleeocapsa sp. in axenic cultures. The growth rate was not affected by concentrations of 0.01 mM or less. This concentration resulted after a lag period in a moderate increase in superoxide dismutase level. After removal of paraquat, the cyanobacterium continued to generate higher levels of superoxide dismutase. There was a lag period of one hour before resumption of normal enzyme activity. Addition of puromycin at concentration of 0.5 mg cm^?3 had no effect on cell survival, but greatly enhanced the sensitivity of the culture to the toxicity of paraquat. The data showed an increase of SOD activity by temperatures above the normal growth temperature level. However, this increase was suppressed by chloramphenicol which revealed that the induction of superoxide dismutase by high temperatures was associated withde novo protein synthesis.     

Enzyme activities ofChlorophyllum molybditis andCortinarius melliolens at different sporophore stages

The intracellular enzyme activities ofChlorophyllum molybditis (Mayerex. Fr.) Massee andCortinarius melliolens Fries were determined at different stages of sporophore maturity. In both mushroom species, total amylase, α-amylase, proteinase, lipase, peroxidase, catalase and polyphenol oxidase activities were increased with sporophore maturity. In contrast, glucose-6-phosphatase activity was higher in the young sporophore than in very young and mature ones. All the enzymes assayed except cellulase and β-amylase showed greater activity in the pilei than in the stipes.C. molybditis showed greater total amylase, α-amylase, cellulase, proteinase, catalase and glucose-6-phosphatase activities thanC. melliolens.