Reversal of the effects of aging in soybean seeds

Accelerated aging predisposed seeds to imbibition injury. Slowing the rate of hydration prevented the loss of germinability due to imbibition injury. Germinability of accelerated aged seeds (50 hours) was increased from 10 to 90% by controlling the rate of imbibition. Slow hydration also prevented seed electrolyte leakage. This may indicate that cell membrane permeability or rupture was a major factor contributing to the loss of germinability after aging.

Reversal of the effects of aging (repair) was accomplished by slowly inbibing and then redrying seeds (priming). This treatment lowered steep water conductivity by a factor of 2 to 5. Priming also increased the per cent germination of low vigor seeds. The mechanism of this reversal was probably metabolic because it depended on temperature, seed moisture, and treatment duration.

Priming doubled the survival of seeds in the accelerated aging vigor test. The `rejuvenation' was accepted as evidence for metabolic repair. Since the `vigor' of seeds was increased by priming, metabolic repair probably included other subcellular components as well as the plasma membrane.

     

The distribution and temporal dynamics of the estuarine macroalgal community of San Francisco Bay

Long-term sampling of intertidal macroalgae along permanently marked transects within San Francisco Bay has shown a marked decline in overall species number along the estuarine gradient from the ocean to the river, presumably as a result of decreasing salinity and a progressive lack of hard substrata in the upstream direction. Green algae penetrated further landward than either brown or red species. Seasonally, macroalgal species diversity is lowest during the winter-spring months when salinity, temperature, and irradiance are at yearly minima. Macroalgal abundance as measured by percent cover was maximum during the late spring near the mouth of the estuary and during late summer towards the head. The seasonal increase in algal abundance is related to increasing salinity, temperature, and light availability to the bottom. The summer increase in irradiance is due to the longer photoperiod, increased frequency of day-time low tides, and reduced levels of suspended sediments. The aperiodic occurrence of algal blooms in San Pablo Bay may be caused by a combination of physical factors which are ultimately associated with the river inflow. A hypothesis based on interannual differences in river inflow and the contribution of phytoplankton to nutrient cycles in the benthos is presented to explain the occurrence of nuisance algal blooms.     

Method for gene replacement in Pseudomonas aeruginosa used in construction of recA mutants: recA-independent instability of alginate production.

The availability of a technique for site-directed mutagenesis by gene replacement provides a powerful tool for genetic analysis in any bacterial species. We report here a general technique for gene replacement in Pseudomonas aeruginosa. Genes on fragments of cloned P. aeruginosa DNA, altered by transposon mutagenesis, can be transduced into a recipient strain and can replace homologous genes in the P. aeruginosa genome. In this study we applied this technique to the construction of recA mutants of P. aeruginosa. A cloned segment of P. aeruginosa FRD1 DNA was isolated which encoded a protein analogous to the recA gene product of Escherichia coli. The P. aeruginosa recA gene was able to complement several defects associated with recA mutation in E. coli. Transposon Tn1 and Tn501 insertions in the cloned recA gene of P. aeruginosa were used to generate chromosomal recA mutants by gene replacement. These recA strains of P. aeruginosa were more sensitive to UV irradiation and methyl methane sulfonate and showed reduced recombination proficiency compared with the wild type. Also examined was the effect of recA mutations on the expression of alginate, a virulence trait. Alginate is a capsulelike polysaccharide associated with certain pulmonary infections, and its expression is typically unstable. The genetic mechanism responsible for the instability of alginate biosynthesis was shown to be recA independent.     

Activity expressed from cloned Anacystis nidulans large and small subunit ribulose bisphosphate carboxylase genes

Summary The ribulose bisphosphate carboxylase/oxygenase (EC4.1.1.39) (RubisCO) large and small subunit genes from Anacystis nidulans have been cloned as a single fragment into M 13mp10 and pEMBL8 and expressed in Escherichia coli. From M 13mp10 a low yield of enzyme with high specific activity was obtained. The molecular weight of the active enzyme was 260 000 Da and of the inactive enzyme approximately 730 000 Da. The small and large subunits cloned separately did not express activity. The RubisCO gene cloned into pEMBL8 expressed activity up to 22 times that from the M 13 cloned RubisCO DNA. The RubisCO protein produced by the pEMBL cloned gene had a normal MW (550 000). Immunoprecipitation and polyacrylamide gel electrophoresis showed the presence of both large and small subunits.     

Effects of transfusion-induced polycythemia on O2 transport during exercise in the dog

An increased hematocrit could enhance peripheral O2 transport during exercise by improving arterial O2 content. Conversely, it could reduce maximal delivery of O2 by limiting cardiac output during exercise or by limiting the distribution of blood flow to peripheral capillaries with high O2 extractions. We studied O2 transport at rest and during graded treadmill exercise in splenectomized tracheostomized dogs at normal hematocrit (38 +/- 3%), and 48 h after transfusion of type-matched donor cells. This procedure increased hematocrit (60 +/- 3%) but also increased blood volume (P less than 0.05). Following transfusion, resting cardiac output (QT) and heart rate were not different. During exercise, QT was significantly lower at each level of O2 consumption (VO2) at high hematocrit (P less than 0.01). A reduction in QT was also seen during polycythemic exercise with hypoxemia produced by breathing 12 or 10% O2 in N2. Despite the reduction in QT, mixed venous PO2 was not lower at high hematocrit, and the increase in base deficit with VO2 was not different from control measurements. O2 delivery (QT X arterial content) was similar at each level of VO2 at both levels of hematocrit, during both normoxic and hypoxic studies. Both systemic and pulmonary arterial pressures were increased at rest after transfusion (P less than 0.05). However, pulmonary and systemic pressures were not higher than control during exercise at high hematocrit. We conclude that a hematocrit of 60% with increased blood volume is not associated with a cardiac limitation of O2 delivery, nor does it interfere with peripheral O2 extraction during exercise in the dog.     

Role of hemoglobin P50 in O2 transport during normoxic and hypoxic exercise in the dog

High hemoglobin affinity for O2 [low PO2 at 50% saturation of hemoglobin (P50)] could degrade exercise performance in normoxia by lowering mean tissue PO2 but could enhance O2 transport in hypoxic exercise by increasing arterial O2 saturation. We measured O2 transport at rest and at graded levels of steady-state exercise in tracheostomized dogs with normal P50 (28.8 +/- 1.8 Torr) and again after P50 was lowered (19.5 +/- 0.7 Torr) by sodium cyanate infusions. Measurements were made during ventilation with room air (RA), 12% O2 in N2, or 10% O2 in N2. Cardiac output (QT) as a function of O2 consumption (VO2) was not altered by low P50 at any inspired O2 fraction (P greater than 0.05). With RA exercise, arterial content (CaO2) and O2 delivery (QT X CaO2) were unchanged at low P50, whereas mixed venous PO2 was reduced at each level of VO2. With exercise in hypoxia, CaO2 and O2 delivery were significantly improved at low P50 (P less than 0.05). Mixed venous PO2 was lower than control during 12% O2 (P less than 0.05) but not different from control during 10% O2 exercise at low P50. Despite a presumed decrease in tissue PO2 during RA and 12% O2 exercise, exercise performance and base excess decline were not significantly worse than control levels. We conclude that, in canine steady-state exercise, hemoglobin P50 is not an important determinant of tissue O2-extraction capacity during normoxia or moderate hypoxia. In extreme hypoxia, low P50 may help to maintain tissue PO2 by enhancing systemic O2 delivery at each level of QT.     

Persistence of Bacillus thuringiensis parasporal crystal insecticidal activity in soil

Spores and parasporal crystals of a Bacillus thuringiensis var. aizawai (H-serotype 7), strain HD137, streptomycin-resistant mutant were added to acidic (pH 5.0) natural and autoclaved soil and incubated at ?0.10 MPa, 25°C. Populations of B. thuringiensis in both soil treatments showed exponential rates of mortality which were represented by linear regression, the loss of viability being greater in natural than autoclaved soil. In natural soil, parasporal crystal insecticidal activity was lost at a complex, nonexponential rate. The initial, rapid decrease of activity gradually slowed, and the level of activity stabilized at 10% of the original inoculum level after 250 days incubation, until the cessation of sampling at >2 years. In autoclaved soil no significant (P > 0.2) loss of parasporal crystal insecticidal activity was detected over the same period, which suggested that soil microorganisms were responsible for the loss of crystal insecticidal activity in the natural, nonsterilized soil. The rate of loss of crystal activity in natural soil correlated well with assay data reported in the literature using Galleria mellonella, which measures the combined activity of spore and crystal. In autoclaved soil correlation was poor, probably due to variability in the bioassay data.     

P. mirabilis RecA protein catalyses cleavage of E. coli LexA protein and the lambda repressor in vitro

Summary The cloned recA ⁺ gene of Proteus mirabilis substitutes for a defective RecA protein in Escherichia coli recA ^– mutants, and restores recombination, repair and phage induction functions to near normal levels. In a previous report, we described the purification and charactrisation of the recombination activities of the P. mirabilis RecA protein (West et al. 1983b). In this paper, we show that the purified protein catalyses the cleavage of both the Escherichia coli LexA protein and the bacteriophage lambda repressor in vitro. These results provide a direct biochemical basis for the interspecies complementation observed in vivo and suggest that P. mirabilis has an SOS regulatory network similar to that of E. coli.     

An RP4-prime containing a 285 kb fragment of rhizobium meliloti pSym megaplasmid: Structural characterization and utilization for genetic studies of symbiotic functions controlled by pSym

Summary We integrated the RP4 plasmid into a selected region of the pSym megaplasmid of Rhizobium meliloti 2011 by homologous recombination between pSym and a cloned fragment of pSym present in the RP4. This cointegrate was used to mobilize into Escherichia coli a Tn5 transposon located on pSym in the vicinity of the site of integration of the RP4. By this technique we obtained a series of RP4-primes that contained large fragments of the pSym megaplasmid and that were most probably generated by IS8 promoted deletions in the RP4-pSym cointegrate. One of them, pGMI42, which carries nitrogenase genes nifD and H as well as nodulation genes, was used for mutagenesis of the corresponding region of pSym after insertion of the Mu prophage into the tet gene. When various (pGMI-42:: Mu)::Tn7 were introduced into R. meliloti 2011 by conjugation, homologous recombination allowed insertion of Tn7 into pSym whereas the pGMI42::Mu was lost due to the suicide effect of Mu. In this way we obtained several symbiotic mutants deficient in either nodulation (Nod^-) or nitrogen fixation (Fix^-) in association with the host plant Medicago sativa.This paper is affectionately dedicated to the memory of Jean-Simon Julliot who initiated and inspired this work and who was killed by an avalanche on February 21, 1982