Bacterial stimulation of adventitious rooting on in vitro cultured slash pine (Pinus elliottii Engelm.) seedling explants.

A bacterium has been isolated that initiates adventitious rooting when co-cultured under in vitro conditions with seedling-produced hypocotylary explants of slash pine (Pinus elliottii). Rooting efficiencies produced through bacterial-explant co-culture range from approximately 15% to greater than 90% over non-treated controls. Explant exposure to the root inducing bacterium has produced no obvious pathology in the regenerated plantlets. Seedling explants rooted by bacterial-explant co-culture have been successfully transitioned to ambient greenhouse conditions.

Multiple shoot production from seedling explants of slash pine (Pinus elliottii, Engelm.).

Hypocotylary explants obtained from 30- to 40-day-old slash pine (Pinus elliottii, Engelm.) seedlings treated with 6-benzylaminopurine produced multiple buds that eventually elongated into axillary shoots. The explants were pulse treated (45-s dip) with 6-benzylaminopurine (22.2, 111, 222 µM) plus a control and cultured on three different basal media containing activated charcoal (0.5% w/v). Hormonal concentration and basal medium were compared for the number and size of axillary shoots induced after 12 and 29 days. The greatest number of axillary shoots was produced by explants that were pulse treated with 111 µM 6-benzylaminopurine and cultured on Gresshoff and Doy medium. The axillary shoots were fewer in number per explant than shoots previously reported resulting from hormonally induced advantitious buds of slash pine, but the axillary shoots developed more rapidly.

Genetic characterization of a mutation that enhances paraquat tolerance in the fern Ceratopteris richardii.

Three nuclear mutations that affect tolerance to the herbicide paraquat have been selected in the fern Ceratopteris richardii. Two of the mutations, pq2 and pq45 are allelic and confer low and moderate tolerance, respectively. A third mutation, pqa6, is not linked to the other two and significantly enhances the level of tolerance when in combination with either pq2 or pq45. The pqa6 mutation does not independently confer tolerance in the absence of the other mutations.

Characterization of (14)C-naphthol uptake in excised root segments of clover (Trifolium pratense L.) and fescue (Festuca arundinaceae Screb.).

An isolated root uptake test (IRUT) was used to characterize the bioaccumulation of (14)C-naphthol in excised root segments obtained from 6-month-old hydroponically grown plants: two varieties of fescue (Festuca arundinaceae Schreb.) and one variety of clover (Trifolium pratense L.). Naphthol uptake rates were directly related to naphthol concentration in the range 0.01 to 0.2 uM. The incubation time required for equilibrium to be reached between naphthol in root tissue and in solution was between 9 and 12 h. Tests using metabolic inhibitors, KCN, NaN3, and DNP, indicated that naphthol uptake may be the result of both passive and active mechanisms. Q10 values for naphthol uptake ranged from 1.05 to 1.16.

Biochemical Studies of Paraquat-Tolerant Mutants of the Fern Ceratopteris richardii.

Enzymes and metabolites associated with mitigation of paraquat toxicity were compared in two paraquat-tolerant mutants and a sensitive wild-type strain of the fern Ceratopteris richardii Brongn. In 21-day-old gametophytes, the specific activities of superoxide dismutase, catalase, peroxidase, glutathione reductase, dehydroascorbate reductase, and ascorbate peroxidase showed no differences that would explain mutant tolerance. Constitutive levels of ascorbate and glutathione also did not differ significantly in the three strains. An experiment testing the inducibility of paraquat tolerance revealed no change in the dose response of mutant or wild type gametophytes after exposure to sublethal concentrations of the herbicide. Uptake of paraquat by whole gametophytes was also equivalent in mutants and wild type. These data suggest that the physiological basis for tolerance in these mutants, unlike several other tolerant biotypes reported, does not lie in the oxygen radical scavenging system, in an inducible stress response, or in a block to whole-plant uptake.

Plantlet regeneration from mature zygotic embryos of eastern white pine (Pinus strobus L.).

Plants were regenerated from whole embryo explants obtained from eastern white pine (Pinus strobus L.) seeds. Embryos were surgically removed and axenically cultured to induce buds in vitro on a modified Murashige and Skoog medium containing various concentrations of 6-benzylaminopurine. Embryos remained on bud induction medium for 21 days and then were transferred to the same basal medium without 6-benzylaminopurine to promote bud development and subsequent shoot elongation. The medium containing 10 µM 6-benzylaminopurine induced the greatest number of shoots per embryo. Rooting was achieved by direct transfer of the shoots to a non-sterile artificial soil mixture followed by multiple treatments with 15 nM 1-naphthaleneacetic acid. Regenerated seedlings are currently growing under greenhouse conditions.

Report on discussion of paper by P. M. Attiwill.

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Report on discussion of paper by P. J. H. Sharpe.

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An in vitro whole plant selection system: paraquat tolerant mutants in the fern Ceratopteris.

A whole plant selection system using the haploid gametophyte generation of the fern Ceratopteris richardii has been developed to select for mutations that confer resistance or tolerance to various selection pressures. The expression of the mutations can be analyzed and characterized in both the haploid gametophyte and diploid sporophyte generations. Genetic analyses are facilitated by the fern's rapid life cycle and the ease of manipulating the gametophyte generation. Selection for tolerance to the herbicide paraquat has yielded two mutants which have an increased tolerance to the herbicide in both the gametophyte and sporophyte generations. Both mutants exhibit single nuclear gene inheritance patterns and appear to be closely linked or allelic.

Thymidine phosphorylation in wheat: analysis of phosphate transfer from ATP to thymidine.

Extracts of wheat (Triticum vulgare Vill. [Triticum aestivum L.] var. Lemhi) seedlings contain thymidine-phosphorylating activity with ATP, ADP, or AMP and nucleotide hydrolase activity (ATP --> --> AMP). The synthesis of [(32)P]dTMP exclusively from [alpha-(32)P]ATP with none detectable from [gamma-(32)P]ATP demonstrates the absence of thymidine kinase and the presence of nucleoside phosphotransferase as the only observable thymidine-phosphorylating enzyme.

Studies of the herbicide paraquat. I. Effects on the cell cycle and DNA synthesis in Vicia faba.

Vicia faba root tips were exposed to concentrations of paraquat ranging from 10-3M to 10-6M. There was a statistically significant inhibition in 3H-thymidine incorporation, and, therefore, presumably of DNA synthesis by all concentrations of paraquat studied. All concentrations of paraquat also had a statistically significant effect on the percentage of cells in division at various hours following paraquat treatment. At 10-3M and 10-4M paraquat there was an almost total inhibition in the number of cells moving from the S period and through the G2 period and into division. Cells treated with 10-6M paraquat, however, did move through the G2 period and into division. A concentration of 10-4M paraquat did not result in an increase in chromosome aberrations.

Large Scale Isolation and Purification of Eyespot Granules from Euglena gracilis var. bacillaris.

Large volumes of eyespot granules were isolated from homogenates of Euglena gracilis Klebs var. bacillaris Pringsheim by flotation centrifugation in a Beckman Ti-15 zonal rotor, and were further purified by centrifugation in a swinging bucket rotor. Examination with the electron microscope showed the eyespot granules to be free from other cellular material. Freezing had no apparent effect on the structure or on the absorption properties of the eyespots. Absorption spectra of pure fractions of eyespot granules free of chloroplast contamination showed the previously reported curves in the range of 360 to 520 nanometers, as well as a peak at 660 to 675 nanometers. The procedure for the large scale isolation of eyespot granules from Euglena gracilis is compared with other methods which have employed conventional centrifugation, and the significance of the use of zonal rotors for isolating large quantities of pure eyespot granules is discussed.