Leaf disc

Leaf discs are floated on a suspension of Agrobacterium ceWs that contain genetically engineered Ti-plasmids. The bacterium infects cells around the edges of the discs, and the T-DNA is transferred to the chromosomes of the infected cells. 

Leaf discs from rapidly expanding V.faba leaves incorporated 14C-labeled palmitic acid into cutin. After removal of the soluble lipids and other materials, the insoluble residue was subjected to LiAlH4 hydrogenolysis and the labeled reduction products of cutin monomers were identified by chromatography as hexadecane-... 

Procedure The leaf disc (< ) 1cm) was placed in a vial containing 1 ml dimethyl sulfoxide and chlorophyll was extracted at 60 °C by incubating for 30 min. After cooling the OD values at 645 and 663 nm were read in a Shimadzu UV 160 spectrophotometer. Chlorophyll a and b content was calculated following the equation used by Amon (1949). The content of chlorophyll a and b was expressed in mg/g dry weight. 

The two remaining of plant water status parameters (water and osmotic potential) reflect free water availability. These can be measured with a Dew Point Microvoltmeter (e.g., the WESCOR HR 33T [Logan, Utah]), but other methods and equipment can be applied. Osmotic potential can be determined from either fresh or frozen plant samples, but water potential requires fresh tissue. The water potential of leaf discs, strips or roots of the same size/weight can be measured with the WESCOR device (for detailed methods see http //www.wescor.com/environmental/index.phtml). Osmotic... 

Leaf discs (10 per treatment) were punched out with a 18-mm-dlameter cork borer Insecticide-treated discs were in a 0.008% (w/v) solution of carbaryl in acetone, and control discs were dipped in acetone alone and allowed to dry. One caterpillar was placed on each disc and held for 24 hr in a 1-oz creamer cup placed in a controlled-environment chamber (25 C, 16/8 hr llgbt/dark)... 

Leaves of Nicotiana tabacum were treated with infective potato-X-virus. About 3 h after infection leaf discs were prepared (10 mm diameter) and incubated on water containing 1 mM sparteine. Controls were incubated on water without alkaloid. At the specified intervals samples (50 discs each) were taken and... 

Table I. Uptake of Am POLLUTION EFFECTS ON PLANT GROWTH sucrose-U-l C by cotton (Acala SJ-1) leaf discs ... 

Figure 2. Effect of ozone on uptake and incorporation of -leucine into protein by cotton cotyledon leaf discs. Plants were exposed to 0.4 ppm Os for 1 hr, 24 hr prior to experiments. Discs were floated on buffer and incubated in -leucine for up to 4 hr and were then transferred to excess cold leucine to chase the incorporated C-leucine for a subsequent 24 hr period. The data show that ozone-treated tissue incorporated more leucine into protein but do not indicate real differential effects on protein hydrolysis.

Figure 3. Logarithmic plot of C-leucine uptake by cotton leaf discs. Data are from Fig. 2.

Figure 4. Time course of uptake rate of 2-deoxyglucose by bean leaf discs following exposure of intact plants to 0.4 ppm ozone. Two standard deviations about the mean are shown. Data of Perchorowicz and Ting (17).

As shown in Table III, there is no significant difference in glucose uptake by cotton leaf discs immediately after exposure, but after 24 hr the uptake has nearly doubled. The percentage distribution of radioactivity in various extraction fractions is approximately the same in control and ozone-treated tissues. CO2 evolution from the added glucose is virtually identical in control and treated tissues. Isotope incorporation into the water soluble fraction, which would include amino acids, organic acids, and soluble carbohydrate, is also quite similar. Some distributional differences in the insoluble fraction are noted between... 

Figure 5. Logarithmic plot of 2-deoxyglucose uptake by bean leaf discs immediately following (day 0) and 24 hr (day 1) after exposure to 0.4 ppm ozone for 1 hr. Data from Perchorowicz and Ting (17).

Figure 6. Time course of tritiated water uptake or release from bean leaf discs previously exposed to 0.5 ppm ozone for 1 hr immediately prior to experiment. Upper Discs were preincubated in 0.2 M mannitol for 2 hr then transferred to tritiated water. Lower Discs were preincubated in tritiated water then transferred to 0.2 M mannitol. Data of Evans and Ting (13).

Salt Transport. The effects of ozone on membrane permeability can also be assessed by estimating salt leakage from treated tissue. In one study, susceptible bean plants were allowed to take up RbCl for 24 hr prior to ozone exposure. After exposure, leaf discs were placed in a desorption solution containing 0.5 M CaSOi and 2 mM KCl and the rate of Rb leakage into the desorption solution was determined. The initial loss was indistinguishable between treated and untreated plants and we assume that it represented exchange from free space. Then, for an extended period, treated tissue exhibited a linear loss of... 

Figure 8. Plot of Rb-K uptake by bean leaf discs as a function of K concentration. Light and dark treatments for control and ozone treated plants are shown. Data of Evans and Ting (15).

Using bean leaf discs fed 1-l C-acetate, we showed that the radioactively labeled diglyceride content of ozonated discs became consistently less, and that this reduction was accompanied by an increase in the radioactivity of ASG (unpublished). 

Similar results were obtained using 12-day-old cucumber leaf discs washed in tap water replicate samples of the discs themselves were taken at short intervals for 60 min. No change in incorporated label with time of washing was observed, and no differences between rates of label loss from soluble pools of control and ozonated (47 2 pphm, 30 min) plants were observed even though the ozonation treatment reduced label influx into the soluble pool by between 35 and 40%. 

Temperature-Dependence of Amino Acid Uptake in Soybean. The uptake of amino acids by plants into the soluble pool can be an energy-requiring process (52-54), as is the support of protein synthesis. We wished to show the temperature-dependence of influx of amino acids into soybean trifoliate leaf discs taken from control and ozonated seedlings in order to confirm this for the present system. Table II shows that amino acid influx during 30 min was reduced by the lowered temperature by about 70%, and protein synthesis by about 85%, when labelling of control discs proceeded at ice bath temperatures. Qualitatively similar reductions occurred in discs taken from ozonated (50 5 pphm,... 

Larvae were individually fed a lima bean leaf disc loaded with a terpene dose of 100 2 pg... 

Several chemicals are known to inhibit photorespiration, the most studied being a-hydroxy-2-pyridine methane sulfonic acid (HPMS Figure 5.6). Early work using leaf discs showed that HPMS caused the accumulation of glycolate in the light and increased carbon fixation at elevated temperatures. 

Due to the sensitivity of the RIA, it is now possible to measure limonin concentration in any part of the plant from embryos to small leaf discs, seed coats or any other desirable site or sample. In this type of analysis or screening study it is possible for a single person to analyze 2000 or more samples per week and it is ironic that "sample taking" is now likely to become the limiting step in any large scale study. 

The third metabolic pathway discovered for degradation of triazine herbicides in plants was first reported by Shimabukuro et al. (1970) and involved conjugation of atrazine with glutathione in corn nutrient uptake, excised leaves, and leaf disc experiments. This new degradation mechanism was postulated to be the primary factor in the... 

L-Cysteine desulfhydrase in leaves of cucurbit plants is a constitutive enzyme whose activity can be enhanced by preincubation of leaf discs with L-cysteine, D-cysteine, or structural analogs of L-cysteine at millimolar concentrations preincubation with cystine does not affect the activity of the enzyme (20.261. Although the stimulation of L-cysteine desulfhydrase activity is... 

Cytokinin activity can be assayed conveniently with several plant tissues, including soybean (54) and tobacco (55,56) callus, radish leaf discs (11,12) and lettuce seed (57). The first two of these are probably the most useful since they are not excessively sensitive to light or temperature (58,59) and reflect both cell enlargement and division (60). 

It should be noted that in most of these studies, especially in those on the barley leaf discs, the metal concentrations applied are significantly higher than those found in the field. It remains, therefore, to be proven that the cadmium effects described occur at the metal concentrations present in intact plants growing in situ on a cadmium-contaminated substrate. 

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