Temperature regulation of oleate desaturase in sunflower (Helianthus annuus L.) seeds

The effect of temperature on oleate desaturation in developing sunflower (Helianthus annuus L.) seeds has been examined. When seeds from plants grown at low (20/10° C, day/night) temperature were transferred for 24 h to 10° C, an increase in the linoleate/oleate ratio in phosphatidylcholine and triacylglycerol was observed, but not when transfer was to 20 or 30° C. The same effect was observed in triacylglycerol, phosphatidylcholine and phosphatidylethanolamine in the newly synthesized lipids after in-vivo incubation with [1-¹⁴C]oleate at 10° C. The microsomal oleoyl phosphatidylcholine desaturase (ODS) activity of the seeds maintained at 10 C was also enhanced. The stimulation was observed after only 3 h in plants grown at high temperature (30/20° C). This effect was inhibited by cycloheximide, implying that the low-temperature stimulation of the ODS activity was caused by the synthesis of new enzyme. As a consequence, seeds from plants grown at low temperature had higher ODS activities and linoleate contents than those grown at high temperature. The microsomal ODS activity of seeds from plants grown at low temperature was dependent on incubation temperature and showed a maximum at 20° C. By contrast, this activity was almost temperature-insensitive in seeds from plants grown at high temperature. These results could explain how temperature regulates the fatty-acid composition in sunflower-seed lipids.Abbreviations DAF days after flowering - ODS oleoyl phosphatidylcholine desaturase - PC phosphatidylcholine - PE phosphatidylethanolamine - TAG triacylglycerol - 181 oleic acid - 182 linoleic acid To whom correspondence should be addressedThanks are due to M.C. Ruiz for skillful technical assistance. This work was supported by a grant from Junta de Andalucia, Spain.     

Leaf area expansion and assimilate production in sunflower (Helianthus annuus L.) growing under low phosphorus conditions

Reductions in leaf area and plant growth as a consequence of phosphorus (P) limitations have been attributed both to direct effects of P shortage on leaf expansion rate and to a reduced production of assimilates required for growth. Canopy assimilation and leaf area expansion are closely interrelated processes. In this work we used experimental and simulation techniques to identify and study their importance in determining leaf area on sunflower (Helianthus annuus L.) growing under P-deficient conditions. Experiment 1 was done outdoors, in Buenos Aires, Argentina, and Experiment 2 in a glasshouse in Wageningen, The Netherlands. In both experiments we studied the effects of soil P addition on leaf appearance, leaf expansion, dry matter accumulation, and leaf photosynthesis of non-water stressed plants grown in pots containing a P-deficient soil. Before sowing the equivalent amounts of 0–600 kg of super phosphate ha^-1 were added to the pots. Phosphorus deficiency delayed leaf appearance increasing the value of the phyllochron (PHY) up to 76%, the rate of leaf area expansion during the quasi-linear phase of leaf expansion (LER) was reduced by up to 74%, with respect to high P plants. Phosphorus deficiency reduced by up to 50% the rate of light saturated photosynthesis per unit of leaf area (AMAX) in recently expanded leaves, while at low levels of leaf insertion in the canopy, AMAX was reduced by up to 85%, when compared to that in high P plants. Phosphorus deficiency also reduced the duration of the quasi-linear phase of leaf expansion by up to eight days. The values of LER were related (r = 0.56, P < 0.05) to the mean concentration of P in all the leaves (Leaves P%) and not to the concentration of P in the individual leaf where LER was determined (r = 0.22, P < 0.4) suggesting that under P deficiency individual leaf expansion was not likely to be regulated by the total P concentration at leaf level. The values of AMAX of individual leaves were related (r = 0.79, P < 0.01) to the concentration of total P in the corresponding leaf (Leaf P%). LER showed a hyperbolic relationship with Leaves P% (R² = 0.94, P < 0.01, n = 13) that saturate at 0.14%. AMAX showed a hyperbolic relationship with Leaf P% (R² = 0.73, P < 0.01, n = 53) that saturated with values of Leaf P% higher than 0.22. A morphogenetic model of leaf area development and growth was developed to quantify the effect of assimilate supply at canopy level on total leaf area expansion, and to study the effects of model parameters on the growth of sunflower plants under P-deficient conditions. With this model we identified the existence of direct effects of P deficiency on individual leaf area expansion. However, we calculated that under mild P stress conditions up to 83% of the reduction in the observed leaf area was explained by the particular effects of P% on the rate of leaf appearance, on the duration of the linear period of leaf expansion, and on the value of AMAX. We also calculated that the effects of P deficiency on the value of AMAX alone, explained up to 41% of the observed reductions in total leaf area between the highest and the intermediate P level in Experiment 2. Possible mechanisms of action of the direct effects of P on individual leaf expansion are discussed in this paper.     

Effects of leaf position and nitrogen supply on the expansion of leaves of field grown sunflower (Helianthus annuus L.)

Leaf growth responses to N supply and leaf position were studied using widely-spaced sunflower plants growing under field conditions. Both N supply (range 0.25 to 11.25 g added N per plant) and leaf position significantly (p=0.001) affected maximum leaf area (LA_max) of target leaves through variations in leaf expansion rate (LER); effects on duration of expansion were small. Specific leaf nitrogen (SLN, g N m^-2) fell quite rapidly during the initial leaf expansion phase (LA < 35% LA_max) but leveled off during the final 65% increase of leaf area. This pattern held across leaf positions and N supply levels. Leaf nitrogen accumulation after 35% LA_max continued up to achievement of LA_max; reductions in the higher SLN characteristic of the initial phase were insufficient to cover the nitrogen requirements for expansion during the final phase. LER in the quasi-linear expansion phase (35 to 100% of LA_max) was strongly associated with SLN above a threshold that varied with leaf position (mean 1.79±0.225 g N m^-2). This contrasts with the response of photosynthesis at high irradiance to SLN, which has previously been shown to have a threshold of 0.3 g N m^-2; in the present work saturation of photosynthetic rate was evident when SLN reached 1.97 g N m^-2. Thus, once the area of a leaf exceeds 35% of LA_max, expansion proceeds provided SLN values are close to the levels required for maximum photosynthesis. However, growth of leaves during the initial expansion phase ensures a minimum production of leaf area even at low N supply levels.     

Systemic acquired resistance in sunflower (Helianthus annuus L.)

Systemic acquired resistance (SAR) to infection by Botrytis cinerea in the leaves of sunflower (Helianthus annuus L.) plants was induced following cotyledon inoculation with B. cinerea or treatment with abiotic inducers. Salicylic acid (SA), benzo-(1,2,3)-thiadiazole-7-carbothioic S-methyl ester (BTH), 2,6-dichloroisonicotinic acid (INA) or EDTA protected sunflower plants against Botrytis infection, that was revealed by a reduction in the number and area of the necrotic lesions in upper leaves after challenge inoculation with the pathogen. SA and BTH were more potent inducers than INA, EDTA or pre-inoculation with the fungus. In addition to resistance to B. cinerea, the upper leaves have also developed resistance to maceration by a mixture of cell wall-degrading enzymes. Calcium nitrate inhibited both the protective effect and the resistance of leaf discs to cell-wall degrading enzymes. All the tested chemicals increased the synthesis and excretion of sunflower phytoalexins--coumarins scopoletin and ayapin and induced the PR-proteins chitinase and 1,3-beta-glucanase, being the inducer effect of each activator correlated with the level of protection against B. cinerea (BTH > SA > INA > EDTA). Thus, SAR induction is mediated by general increase of plant defence responses. This is the first report on SAR in sunflower.     

Acyl-acyl carrier protein thioesterase activity from sunflower (Helianthus annuus L.) seeds

During sunflower (Helianthus annuus L.) seed formation there was an active period of lipid biosynthesis between 12 and 28 days after flowering (DAF). The maximum in-vitro acyl-acyl carrier protein (ACP) thioesterase activities (EC 3.1.2.14) were found at 15 DAF, preceding the largest accumulation of lipid in the seed. Data from the apparent kinetic parameters, V _max and K _m, from seeds of 15 and 30 DAF, showed that changes in acyl-ACP thioesterase activity are not only quantitative, but also qualitative, since, although the preferred substrate was always oleoyl-ACP, the affinity for palmitoyl-ACP decreased, whereas that for stearoyl-ACP increased with seed maturation. Bisubstrate assays carried out at 30 DAF seemed to indicate that the total activity found in mature seeds is due to a single enzyme with 100/75/15 affinity for oleoyl-ACP/stearoyl-ACP/palmitoyl-ACP. In contrast, at 15 DAF, enzymatic data together with partial sequences from cDNAs indicated the presence of at least two enzymes with different properties, a FatA-like thioesterase, with a high affinity for oleoyl-ACP, plus a FatB-like enzyme, with preference for long-chain saturated fatty acids, both being expressed during the active lipid biosynthesis period. Competition assays carried out with CAS-5, a mutant with a higher content of palmitic acid in the seed oil, indicated that a modified FatA-type thioesterase is involved in the mutant phenotype. Received: 17 December 1999 / Accepted: 25 February 2000     

Light-associated nitrogen distribution profile in flowering canopies of sunflower (Helianthus annuus L.) altered during grain growth

In vegetative canopies of many species, the vertical gradient of lamina nitrogen concentration (NW) parallels the profile of light distribution in such a way that the actual nitrogen partitioning approaches the optimum pattern for canopy photosynthesis. This paper evaluates the hypothesis that a strong sink for nitrogen, viz. growing grain, affects the pattern of lamina nitrogen distribution usually described for vegetative canopies. The light and NW profiles of sunflower (Helianthus annuus L.) crops were characterised from anthesis to physiological maturity. The factorial combination of two plant populations (2.4 and 4.8 plants m^–2) and two levels of nitrogen supply (0 and 5 g N m^–2) were the sources of variation for NW and light profiles. Before the onset of nitrogen accumulation in grain, the pattern of NW was similar to that described for other species and it was related to the distribution of light in the canopy. Important changes in the profile of NW occurred during grain filling that were unrelated to the light regime. Nitrogen was mobilised from leaves in all positions in the canopy and the rate of NW change was greater in leaves closer to the grain, which were also the leaves where nitrogen was more concentrated. It is concluded that the physiological mechanisms involved in determining the distribution of leaf nitrogen in vegetative canopies do not apply to sunflower during grain filling.     

A shoot regeneration protocol effective on diverse genotypes of sunflower (Helianthus annuus L.)

Summary We describe a protocol, and several experiments that helped lead to its development, for sunflower regeneration. Important factors for sunflower regeneration were explant age, cytokinin type and concentration, basal medium, and explant source. We could not induce shoot regeneration from the explants derived from mature tissues including leaf, petiole, and stem. However, use of juvenile explants such as embryo meristem and primordial leaf tissues allowed routine regeneration of 17 different sunflower genotypes. High frequency of shoot regeneration was achieved with these explants taken from seedlings up to 5 d after germination. Explant age was less critical for embryo meristem explants than for primordial leaf tissues. Of the four basal media tested, MS and B5 media produced higher shoot-regeneration frequencies than did Anderson and woody plant media. The highest shoot-regeneration frequency was obtained with MS medium supplemented with 2 μM BA and without auxin. Addition of 1 μM naphthalene-acetic acid to the medium significantly reduced both the percentage of explants producing shoots and average number of shoots per explant. Regenerated shoots were grown to maturity in a greenhouse.     

Screening of plant growth promoting Rhizobacteria isolated from sunflower (Helianthus annuus L.)

Background and Aims

This study was aimed at assessing the diversity of putatively diazotrophic rhizobacteria associated with sunflower (Helianthus annuus L.) cropped in the south of Brazil, and to examine key plant growth promotion (PGP) characteristics of the isolates for the purposes of increasing plant productivity.

Methods

299 strains were isolated from the roots and rhizosphere of sunflower cultivated in five different areas using N-free media. 16S rDNA PCR-RFLP and 16S rRNA partial sequencing were used for identification and the Shannon index was used to evaluate bacterial diversity. Production of siderophores and indolic compounds (ICs), as well phosphate solubilization activities of each isolate were also evaluated in vitro. On the basis of multiple PGP activities, eight isolates were selected and tested for their N-fixation ability, and their capacity as potential PGPR on sunflower plants was also assessed.

Results

All except three Gram-positive strains (phylum Actinobacteria) belonged to the Gram-negative Proteobacteria subgroups [Gamma (167), Beta (78), and Alpha (50)] and the family Flavobacteriaceae (1)]. Shannon indexes ranged from 0.96 to 2.13 between the five sampling sites. Enterobacter and Burkholderia were the predominant genera isolated from roots and rhizosphere, respectively. Producers of siderophores and ICs were widely found amongst the isolates, but only 19.8% of them solubilized phosphate. About 8% of the isolates exhibited all three PGP traits, and these mostly belonged to the genus Burkholderia. Four isolates were able to stimulate the growth of sunflower plants under gnotobiotic conditions.

Conclusions

Enterobacter and Burkholderia were the dominant rhizospheric bacterial genera associated with sunflower plants. Inoculation with isolates belonging to the genera Achromobacter, Chryseobacterium, Azospirillum, and Burkholderia had a stimulatory effect on plant growth.     

Enzymatic characterisation of high-palmitic acid sunflower (Helianthus annuus L.) mutants

Two high-palmitic acid sunflower (Helianthus annuus L.) mutants, CAS-5 and CAS-12, have been biochemically characterised. The enzymatic activities found to be responsible for the mutant characteristics are β-keto-acyl-acyl carrier protein synthetase II (KASII; EC 2.3.1.41) and acyl-acyl carrier protein thioesterase (EC 3.1.2.14). Our data suggest that the high-palmitic acid phenotype observed in both mutant lines is due to the combined effect of a lower KASII activity and a higher thioesterase activity with respect to palmitoyl-acyl carrier protein (16:0-ACP). The level of the latter enzyme appeared to be insufficient to hydrolyse the produced 16:0-ACP completely. As a consequence of this, three new fatty acids appear: palmitoleic acid (16:1 Δ9), asclepic acid (18:1 Δ11), and palmitolinoleic acid (16:2 Δ9 Δ12). These fatty acids should be synthesised from palmitoyl-ACP or a derivative by the action of the stearoyl-ACP desaturase, fatty acid synthetase II and oleoyl-phosphatidylcholine desaturase, respectively. Received: 11 July 1998 / Accepted: 10 October 1998     

Prolyl iminopeptidase from seeds of sunflower (Helianthus annuus L.)

Prolyl iminopeptidase from sunflower seed (Helianthus annuus L.) was purified to molecular homogeneity. It is a 105-kDa heterodimer consisting of two subunits: 53 and 55 kDa. It has pI of 6.2 and optimal activity at pH 8.0–8.5 and 45–50°C. The inhibitory analysis was inconclusive about its catalytic machinery, as a significant degree of modification was not observed with any of the used diagnostic inhibitors. Its specificity is restricted to removal of N-terminal prolyl residues.     

EDTA-enhanced lead phytoremediation in sunflower (Helianthus annuus L.) hydroponic culture

The aim of the present study was to investigate the capability of Sunflower (Helianthus annuus L.) to tolerate and accumulate high amount of lead (Pb) and propose it for soil phytoremediation. To this regard, plants were grown in hydroponics and treated with different Pb concentrations (10 to 160 ??M) and a fixed concentration (500 ??M) EDTA (ethylene diamine tetra acetic acid) for 14 and 28 days (d). Effects on total biomass production, photosynthetic pigments and protein contents as well as the quantities of non protein thiols (NP-SH), glutathione (GSH), phytochelatins (PCs) and activity of glutathione reductase (GR) were estimated. Results revealed that roots (575 ??g g^?1 DW) and shoots (135 ??g g^?1 DW) accumulated Pb after 28 d of exposure, however, addition of EDTA enhanced the Pb accumulation in roots (645 ??g g^?1 DW) and shoots (255 ??g g^?1 DW ). Exposure of Pb (28 d) registered a significant (P?<?0.05) reduction in growth parameters and induction of phytochelatins (P?<?0.05; r?=?0.26) plus some of the important antioxidants (P?<?0.05; r?=?0.42), which were positively correlated to metal accumulation. Sunflower exposed at 40 ??M of Pb for 28 d synthesized higher quantity of PC₂ (18.5 fold) and PC₃ (10.5 fold), as compared to control. However, the results showed that addition of EDTA resulted in low toxicity compared to Pb alone. These data support the capability of H. annuus L. to accumulate and tolerate significant quantity of Pb and its utility for phytoremediation. This is because of the plant has the capacity to combat metal induced oxidative stress via significant synthesis of NP-SH, GSH and high activity of GR, as it would provide sufficient GSH not only for PCs synthesis but also for antioxidant function.     

Cavitation fatigue and its reversal in sunflower (Helianthus annuus L.)

"Cavitation fatigue" is the increased susceptibility of a xylem conduit to cavitation as a result of its prior cavitation. It was investigated whether cavitation fatigue induced in vivo could be repaired in intact plants. Sunflowers (Helianthus annuus L.) were subjected to soil drought in the greenhouse. Native embolism and vulnerability to cavitation was measured in well-watered controls and after 5 d and 10 d of controlled drought. A dramatic cavitation fatigue was observed where droughted xylem that was refilled in the laboratory developed up to 60 PLC (percentage loss of hydraulic conductivity) at -1 MPa versus only 5.2 PLC in non-droughted controls. Rewatered plants showed the complete reversal of cavitation fatigue over 4 d. Reversal of fatigue was correlated with the refilling of embolized vessels in the intact plants (r(2)=0.91, P<0.01), suggesting that xylem transport to fatigued vessels was required for their repair. The in vivo reversal of fatigue was partially duplicated in excised stem segments by perfusing them with root exudates from droughted (DR) and well-watered (WW) plants. The DR exudate had a greater effect, and this was associated with a greater pH in the DR versus WW saps, but there was no difference in total cation concentration. Perfusions with 2 mM CaCl(2) and KCl solutions also partially reversed cavitation fatigue as opposed to no effect with deionized water, suggesting a role of ions in addition to a pH effect. It is suspected that fatigue is caused by stretching and partial disruption of linkages between cellulose microfibrils in inter-conduit pit membranes during air seeding, and that the reversal of fatigue involves restoring these linkages by ingredients in xylem sap.     

Photosynthesis and chlorophyll fluorescence in sunflower (Helianthus annuus L.) leaves as affected by phosphorus nutrition

The effects of phosphate concentration on plant growth and photosyntheticprocesses in primary leaves of young sunflower (Helianthus annuusL.) plants were examined. Plants were grown for 3 weeks on half-strengthHoagland's solution containing 0, 0.1, 0.5, 1.0, and 3.0 molm^–3 orthophosphate (Pi). It was shown that optimal photosynthesisand the highest light utilization capacity were achieved at0.5 mol m^–3 Pi in the growth medium, which was in goodagreement with the maximum content of organic phosphorus inthe leaves. Low phosphate in the medium inhibited plant growthrate. Phosphate deficiency appreciably decreased photosyntheticoxygen evolution by leaves, the efficiency of photosystem two(PSII) photochemistry and quantum efficiency of PSII electrontransport. High oxidation state of PSII primary electron acceptorQA, at 0.1 mol m^–3 Pi, however, indicates that photosyntheticelectron transport through PSII did not limit photosynthesisin Pi-deficient leaves. The results indicate that diminishedphotosynthesis under sub- and supra-optimal Pi was caused mainlyby a reduced efficiency of ribulose 1, 5-bisphosphate (RuBP)regeneration at high light intensities. These results suggestthat, under non-limiting C0₂ and irradiance, photosynthesisof the first pair of leaves could be diminished by both sub-and supra-optimal phosphorus nutrition of sunflower plants. Key words: Helianthus annuus L, phosphate nutrition, photosynthesis, photochemical efficiency     

Transformation of sunflower (Helianthus annuus L.): a reliable protocol

Summary A reliable protocol for the transformation of cultivated sunflower (Helianthus annuus L.) has been established, based on microprojectile bombardment of half shoot apices in combination with Agrobacterium tumefaciens coculture. Transgenic shoots have been obtained from 5 inbred lines, although transformation efficiencies varied with the genotype. Plants expressing the transgenes could be recovered from up to 7% of the explants. A minority of plants was shown to be chimaeric for expression of ß-glucuronidase activity while most appeared to be uniformly transformed. Genetic segregation was 31 for both ß-glucuronidase and neomycine phospho transferase in some plants, indicating that the respective mother plants were uniformly transformed. Integration of the foreign genes was also shown by Southern analysis.Abbreviations BAP benzyl amino purine - EDTA ethylene diamine tetraacetic acid - GUS ß-glucuronidase - npt II neomycine phospho-transferase II     

Nuclear DNA content in differentiated tissues of sunflower (Helianthus annuus L.)

Summary Scanning cytophotometry following Feulgen-staining was used to determine nuclear DNA content in many differentiated tissues of nine cultivars, hybrids or selfed lines ofHelianthus annuus. Apart from such ephemeral tissues as endosperm and anther tapetum, it was found that tissue differentiation in sunflower occurs in the diploid condition, cells being arrested in the DNA presynthetic phase (G₁). In certain cases, however, the nuclear DNA content of differentiated G₁ cells does not exactly match the 2C DNA content found in meristematic cells, but may be either higher or lower. In endosperm and anther tapetum cells, nuclear DNA content may be as high as 24 C and 32 C, respectively. Cytological and autoradiographic analyses after³H-thymidine incorporation reveal that polyploidy in the tapetal cells is due to chromosome endoreduplication. No detectable difference between male-fertile and male-sterile plants exists as far as occurrence and level of cell polyploidy are concerned. The results are discussed in the context of previous investigations on the nuclear condition of differentiatedHelianthus annuus tissue.     

Comparative genetic analysis of quantitative traits in sunflower (Helianthus annuus L.)

One hundred and fifty F₂–F₃ families from a cross between two inbred sunflower lines FU and PAZ2 were used to map quantitative trait loci (QTL) for resistance to white rot (Sclerotinia sclerotiorum) attacks of terminal buds and capitula, and black stem (Phoma macdonaldii). A genetic linkage map of 18 linkage groups with 216 molecular markers spanning 1,937 cM was constructed. Disease resistances were measured in field experiments for S. sclerotiorum and under controlled conditions for P. macdonaldii. For resistance to S. sclerotiorum terminal bud attack, seven QTL were identified, each explaining less than 10% of phenotypic variance. For capitulum attack by this parasite, there were four QTL (each explaining up to 20% of variation) and for P. macdonaldii resistance, four QTL were identified, each having effects of up to 16%. The S. sclerotiorum capitulum resistance QTL were compared with those reported previously and it was concluded that resistance to this disease is governed by a considerable number of QTL, located on almost all the sunflower linkage groups.     

Oleate from triacylglycerols is desaturated in cold-induced developing sunflower (Helianthus annuus L.) seeds

For the first time, an active fatty-acid metabolism is indicated for triacylglycerols (TAG) of developing sunflower (Helianthus annuus L.) seeds. When the developing seeds were transferred to low temperature, the total amount of oleate found in TAG decreased as that of linoleate increased, while the contents of total lipids and TAG remained unchanged. These results suggest that oleate from TAG was used for desaturation. This occurred first in microsomal TAG, but after a long cold period it was observed mainly in the oil-body fraction. Thesn-2 position of TAG was preferentially enriched in linoleate. Apparently, more linoleate than necesary for the maintenance of membrane fluidity was synthesized at the expense of TAG oleate.     

Effect of field limestone applications on cadmium content of sunflower (Helianthus annuus L.) leaves and kernels

Nonoilseed sunflower (Helianthus annuus L.) is naturally higher in cadmium (Cd) than many other grain crops. Because raising soil pH usually depresses Cd uptake by most species, a study was designed to determine if application of agricultural limestone to neutralize soil acidity would decrease Cd uptake by sunflower plants grown on different soils in the production area of North Dakota. The field experiments were conducted at 3 locations in 1991 and 2 locations in 1992. At each site, limestone was applied to bring soil pH to 6.5–7.0, or an additional 45 Mg ha^-1 more limestone was applied, and these two treatments were compared to no-lime control. Commercial nonoilseed hybrid 954 was planted in these experiments. The rapid short-term lime-soil reaction occurred in first 12 weeks following limestone application. Mean kernel Cd concentration for each treatment varied from 0.35 to 1.45 mg kg^-1 DW in the first year of the experiments, and from 0.37 to 1.23 mg kg^-1 DW in the experiments of 1992 across all locations. Large variations in kernel Cd levels between locations were obtained. There were no significant differences among control and limestone treatments for kernel Cd, seedling leaf Cd and diagnostic leaf Cd within each location, respectively. In regression analysis, we found that kernel Cd level correlated with diagnostic leaf Cd concentration in each treatment, but poor correlations were obtained among other variables. These results indicated that limestone application did not reduce Cd uptake and transfer to kernels of sunflower, in contrast with most species studied.     

Transformation of sunflower (Helianthus annuus L.) following wounding with glass beads

A procedure was developed for transformation of Helianthus annuus (sunflower) using Agrobacterium tumefaciens. Cotyledons were removed from young seedlings, and the remaining tissue was uniformly wounded by shaking with glass beads. The wounded tissue was then co-cultivated with a hypervirulent strain of Agrobacterium tumefaciens harboring the binary plasmid pCNL56. Minimal use of defined medium was required, and no callus was observed. The polymerase chain reaction (PCR) followed by DNA hybridization demonstrated the presence of gusA DNA from pCNL56 in total leaf DNA of 6 primary transformants and 2 progeny plants. No Agrobacterium DNA was detected in total DNA from transformed sunflower leaves that was amplified with primers specific to the miaA chromosomal gene of Agrobacterium. Foreign DNA was also detected in the next generation. -Glucuronidase (GUS) activity was demonstrated for 5 of the T₂ transgenic plants. Grafting was used to increase the number of seeds present on plants that had undergone tissue culture manipulations.Abbreviations PCR polymerase chain reaction - EMBL European Molecular Biology Laboratory - M U 7-hydroxy-4-methylumbelliferone - GUS -Glucuronidase Disclaimer: Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products that may also be suitable.     

The influence of ancymidol on morphology,anatomy, and chlorophyll levels in developing and mature Helianthus annuus leaves

Ancymidol foliar spray at 132 mg·liter^–1 a.i. modified leaf anatomy of developing Helianthus annuus L. Mammoth Russian leaves, but not of mature leaves. Ancymidol was effective in retarding plant growth when applied at the young seedling or at a more mature stage of growth. Ancymidol increased leaf weight per unit area and chlorophyll content on an area and unit weight basis, regardless of stage of leaf development. Total chlorophyll per leaf was also increased in mature leaves. Thus, darker green foliage due to increased chlorophyll content and modified leaf anatomy responses were determined to be independent effects.Texas Agricultural Experiment Station paper no. 22927.