Pumpkin cotyledons

Leaf discs have commonly been used for bioassays to determine if herbicides inhibit photosynthesis (Table 16.2). The simplest leaf-disc bioassay uses small discs cut from fully expanded cucumber or pumpkin cotyledons, floated in the light on a phosphate buffered medium containing suspected photosynthesis inhibitors.115 Qualitatively, if photosynthesis is inhibited, the leaf disc sinks. There are several variations of this method that can provide quantitative data. Evolution of O2 in the test solution can be measured with an oxygen electrode, CO2 induced pH changes colorimetrically determined with bromothymol-blue, or electrolyte leakage monitored with a conductivity meter. Leaf strips, algae, isolated chloroplasts, and duckweed (Lemna minor) have been used as test subjects. Although the bioassays presented in Table 16.2 are fairly easy to perform, few allelochemicals have been tested as possible inhibitors of photosynthesis. Many... 

Evidence for a separate cytoplasmic GDH was, however, presented by Chou and Splittstoesser (1972) who showed that NAD-linked GDH enzymes in mitochondrial and supernatant fractions of pumpkin cotyledons differed in certain kinetic properties, and that supernatant and mitochondrial fractions possessed different numbers of GDH isoenzymes, as determined by polyacrylamide gel electrophoresis. 

Ca " ", Mn, and Cu " activated the amination reaction of the soluble enzyme from pumpkin cotyledons, but had no effect on the particulate enzyme (Chou and Splittstoesser, 1972). While both Zn + and Ca have been found to reverse inactivation of higher plant GDH enzymes by EDTA (Yamasaki and Suzuki, 1969), Zn + were found to inhibit the amination reaction of enzymes from pea seedlings (Pahlich and Hoffman, 1975) and pumpkin cotyledons (Chou and Splittstoesser, 1972). 

Fig. 2. Effect of phytohormones on phosphorylation of ribosomal protein S< in detached pumpkin cotyledons. The autoradiograms of one-dimensional SDS gel electrophoresis of ribosomal proteins isolated from pumpkin cotyledons. Cotyledons were incubated for 6 h in water a and in solution of b ABA (10 M) c BA (5x lO M) or d BA-I-ABA in the same concentrations respectively. Discs from cotyledons were then labeled for 2 h with P (50 jnCi/ml). and isolated ribosomal proteins were subjected to electrophoresis [26]...

MOLECULAR CHARACTERIZATION OF ACONITASE IN ETIOLATED PUMPKIN COTYLEDONS. 

In the present study, we isolated and characterized pumpkin cytosolic aconitase cDNA. Fluorescent microscopic observation of the etiolated pumpkin cotyledons using an antibody raised against the aconitase expressed in E. coli clearly showed that no glyoxysomal aconitase exists in etiolated pumpkin cotyledons. 

A X.gtll cDNA library prepared from RNA of 4 day-old etiolated pumpkin cotyledons was screened using an affinity purified antibody against pumpkin aconitase [2]. The aconitase cDNA of 3154 bp was obtained by the combination of immunoscreening and plaque hybridization technique. The aconitase cDNA contains an open reading frame of2694 bp capable of encoding a 98 kD protein. 

Cell isolated from pumpkin cotyledons was stained with anti-aconitase antibody and visualized with FITC-conjugated secondary antibody. Arrow shows one of the small fluorescent spots. Bar indicates 10pm. 

De Beilis L, Tsugeki R, Alpi A. Nishimura M. Purification and characterization of aconitase isoforms from etiolated pumpkin cotyledons. Physiol Plant 1993 88 485-492. 

De Beilis L, Hayashi M, Biagi PP, Hara-Nishimura I, Alpi A. Nishimura M. Immunological analysis of aconitase in pumpkin cotyledons the absence of aconitase in glyoxysomes. Physiol Plant 1994 90 757-762. 

The soluble 2-oxoglutarate-dependent dioxygenases that catalyse the later steps of the pathway (part 3) include principally the 20-oxidase, 2 3- and 3p-hydroxylases. They couple GA oxidation with the decarboxylation of 2-oxoglutarate to succinate and require Fe " and ascorbate for maximum catalytic rates. Several of these enzymes have been purified to different degrees [reviewed in 67,68], but only GA, 20-oxidase from pumpkin endosperm has been obtained from plant tissues in a pure form [69], allowing a cDNA clone for this enzyme to be isolated from the immature cotyledons [70]. It contained an open reading frame encoding a protein of 43.3 KDa and, after expression of the cDNA in... 

In 1969, Knypl (10) found that an onium compound, chlorme-quat [(2-chloroethyl)-trimethylammonium chloride], caused all-trans-lycopene accumulation in detached pumpkin (Curcubita pepo) cotyledons. However, no such response was observed when chlorme-quat was applied to other plant tissues (11). A major step forward was observed in 1970 with the discovery that another onium compound, 2-(4-chlorophenylthio)-triethylammonium chloride (CPTA), can cause the accumulation of lycopene in a wide array of plant tissues and microorganisms (11). 

Bean cotyledons Spinach leaves Spinach leaves Spinach leaves Bean embryos Pea embryos Pumpkin endosperm... 

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