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PubMed2000

CDNA cloning of radish (Raphanus sativus) myrosinase and tissue-specific expression in root.

Two cDNA clones of myrosinase (thioglucoside glucohydrolase, EC 3.2.3.1) were isolated from radish (Raphanus sativus) seedlings. Both clones were identified as MB (B type myrosinase) from their sequence homology at the amino acid level to MBs cloned from other Brassicaceae species. The tissue distribution of gene expression and enzyme activity of myrosinase corresponded well to the site of glucosinolate accumulation in different tissues of radish. The myrosinase-glucosinolate system was localized in the cotyledons in the seedlings and in the peel of the root in the mature plant. Tissue printing analysis showed that myrosinase mRNA and activity were localized in the epidermis and vascular cambium that were present in the peripheral part of the root but few signals were detected in the parenchyma inside of the vascular cambium. Since the myrosinase-glucosinolate system is known to be a defense system in higher plants, the localization of the myrosinase-glucosinolate system in the peel of the root may act to protect the sink organ from the attack of herbivores or pathogens in soil.

Amino Acid Sequence↗
PubMed1992

alpha-l-Arabinofuranosidase from Radish (Raphanus sativus L.) Seeds.

An alpha-l-arabinofuranosidase has been purified 1043-fold from radish (Raphanus sativus L.) seeds. The purified enzyme was a homogeneous glycoprotein consisting of a single polypeptide with an apparent molecular weight of 64,000 and an isoelectric point value of 4.7, as evidenced by denaturing gel electrophoresis and reversed-phase or size-exclusion high-performance liquid chromatography and isoelectric focusing. The enzyme characteristically catalyzes the hydrolysis of p-nitrophenyl alpha-l-arabinofuranoside and p-nitrophenyl beta-d-xylopyranoside in a constant ratio (3:1) of the initial velocities at pH 4.5, whereas the corresponding alpha-l-arabinopyranoside and beta-d-xylofuranoside are unsusceptible. The following evidence was provided to support that a single enzyme with one catalytic site was responsible for the specificity: (a) high purity of the enzyme preparation, (b) an invariable ratio of the activities toward the two substrates throughout the purification steps, (c) a parallelism of the activities in activation with bovine serum albumin and in heat inactivation of the enzyme as well as in the inhibition with heavy metal ions and sugars such as Hg(2+), Ag(+), l-arabino-(1-->4)-lactone, and d-xylose, and (d) results of the mixed substrate kinetic analysis using the two substrates. The enzyme was shown to split off alpha-l-arabinofuranosyl residues in sugar beet arabinan, soybean arabinan-4-galactan, and radish seed and leaf arabinogalactan proteins. Arabinose and xylose were released by the action of the enzyme on oat-spelt xylan. Synergistic action of alpha-l-arabinofuranosidase and beta-d-galactosidase on radish seed arabinogalactan protein resulted in the extensive degradation of the carbohydrate moiety.

Journal Article↗
PubMed1971

Red and far red effects on phenylalanine ammonia-lyase in raphanus and sinapis seedlings do not correlate with phytochrome spectrophotometry.

In seedlings of Raphanus sativus (radish) and Sinapis alba (mustard), irradiation for 6 hours with far red light significantly increases the extractable activity of phenylalanine ammonia-lyase by the end of the light period. A schedule of 10 minutes red light-110 minutes darkness-10 minutes red-110 minutes darkness-10 minutes red-110 minutes darkness has no effect as compared to dark controls. However, the red light program maintains a level of far red-absorbing phytochrome always measurable by in vivo spectrophotometry during the 6-hour experimental period. We conclude that the far red effect on this enzyme and for this specific material cannot be explained solely by formation and maintenance of far red-absorbing phytochrome.

Journal Article↗
PubMed1972

In Vivo and In Vitro Studies on gamma-Aminobutyric Acid Metabolism with the Radish Plant (Raphanus sativus, L.).

Labeled glutamate was rapidly converted to gamma-aminobutyrate in intact, excised radish (Raphanus sativus L., var. Champion) leaves. Labeled gamma-aminobutyrate was metabolized via succinate and the Krebs cycle and was not carboxylated to form glutamate. Administration of carbon-14 and tritium-labeled succinate indicated that less than 10% of the gamma-aminobutyrate formation occurs by amination of succinic semialdehyde. Therefore, most gamma-aminobutyrate formation must be via glutamate decarboxylation.Radish leaf extracts were more active in catalyzing transamination between gamma-aminobutyrate and pyruvate than that between gamma-aminobutyrate and alpha-ketoglutarate. Glutamate decarboxylase was approximately 20 times more active than gamma-aminobutyrate: pyruvate transaminase. Succinic semialdehyde dehydrogenase was found in the extracts, and NAD was much more active as a hydrogen acceptor than NADP. No reduction of succinate to succinic semialdehyde by the NAD-linked dehydrogenase could be demonstrated. The following pH optima were determined: glutamate decarboxylase, 5.9; gamma-aminobutyrate: pyruvate transaminase, 8.9; succinic semialdehyde: NAD dehydrogenase, about 9.0.

Journal Article↗
PubMed1974

Adenosine Phosphates in Germinating Radish (Raphanus sativus L.) Seeds.

Changes in concentrations of adenosine phosphates (AMP, ADP, and ATP), oxygen utilization, and fresh weights were measured during the first 48 hours after imbibition of water by quiescent radish seeds (Raphanus sativus L.) at 22.5 C. The changes in ATP concentrations, oxygen utilization, and fresh weights followed a triphasic time course, characterized by a rapid initial increase, which extended from 0 to approximately 1.5 hours, a lag phase from 1.5 to 16 hours, and a sharp linear increase from 16 to 48 hours. In unimbibed seeds, the concentrations of ATP, ADP, and AMP were <0.1, 0.9, and 2.2 nmoles/seed, respectively. After imbibition of water by the quiescent seeds, for 1 hour, the ATP concentration had increased to 2.5, and ADP and AMP concentrations had decreased to 0.3 and 0.1 nmole/seed, respectively. These early changes occurred also in seeds maintained under anaerobic conditions (argon), or when treated with either 5 mm fluoroacetate, or 5 mm iodoacetate. The concentrations of ADP and AMP did not change significantly from 1 to 48 hours. The termination of the lag phase at 16 hours correlated with radicle emergence. Cell division in the radicles was initiated at approximately 28 hours. ATP concentrations in seeds maintained under argon or treated with fluoroacetate remained relatively constant from approximately 2 to 48 hours. In contrast, the ATP concentration of iodoacetate-treated seeds decreased curvilinearly from 4 to 48 hours. Oxidative phosphorylation was estimated to have contributed 15, 20, and 65% of the pool ATP at 1.5, 16, and 48 hours, respectively.

Journal Article↗
PubMed1984

Characterization of Radish (Raphanus sativus) Storage Proteins.

Radish (Raphanus sativus cv Rond rose à bout blanc Vilmorin) seeds, as other cruciferae oil seeds, contain two major types of storage protein aggregates which can be separated by gel filtration into 12 and 1.7 Svedberg fractions. These two fractions have been characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, amino acid composition, and two bidimensional gel electrophoresis systems. These results were compared with those obtained with rapeseed storage proteins. Radish 12 Svedberg particles are made of a series of nine major polypeptides ranging from 33 to 30 kilodaltons. These polypeptides present charge heterogeneity. The 12 Svedberg particle is made of six subunits approximately 55 kilodaltons. Each subunit is a couple of two polypeptides linked by a disulfide bridge. The 1.7 Svedberg particle has a simpler composition. It is made of two polypeptides of 10 and 12 kilodaltons and smaller peptides of approximately 7 kilodaltons. Twelve and 1.7 Svedberg particles also differ in their amino acid composition, the 1.7 Svedberg being particularly rich in glutamic acid and proline. Its components are basic. The organization of the rapeseed storage protein is similar but more complex.

Journal Article↗
PubMed1984

Methylation Pattern of Radish (Raphanus sativus) Nuclear Ribosomal RNA Genes.

The methylation pattern of radish Raphanus sativus nuclear rDNA has been investigated using the Hpa II, Msp I, and Hha I restriction enzymes. The presence of numerous target sites for these enzymes has been shown using cloned rDNA fragments. A large fraction of the numerous rDNA units are heavily methylated, being completely resistant to Hpa II and Hpa I. However, specific sites are constantly available in another fraction of the units and are therefore unmethylated. The use of different probes allowed us to demonstrate that hypomethylated sites are present in different regions. Major hypomethylated Hha I sites have been mapped in the 5' portion of 25S rRNA coding sequence. Among the hypomethylated fraction, different methylation patterns coexist. It has been possible to demonstrate that methylation patterns are specific for particular units. The Hha I pattern of rDNA in tissues of different developmental stages was analyzed. Evidence for possible tissue specific differences in the methylation pattern is reported.

Journal Article↗
PubMed1988

Changes in the Levels of Calmodulin and of a Calmodulin Inhibitor in the Early Phases of Radish (Raphanus sativus L.) Seed Germination: Effects of Aba and Fusicoccin.

An inhibitor of Ca(2+)-calmodulin (Cam)-dependent brain phosphodiesterase was present in the soluble fraction of embryo axes from ungerminated radish (Raphanus sativus L.) seeds. This inhibitor is a Ca(2+)-dependent, Cam-binding protein; in fact: (a) its effect was strongly reduced by treatment with proteases; (b) the inhibition was counteracted by Cam but not by Ca(2+); (c) on gel filtration in the presence of Ca(2+), Cam co-chromatographed with the inhibitor. The inhibitor is heat stable and positively charged at pH 7.5. During early phases of germination, the fresh weight and the levels of DNA and RNA of embryo axes increased, the level of the inhibitor decreased, and the level of Cam increased. Abscisic acid (ABA) inhibited germination, the decrease of inhibitor, and the increase of Cam. Fusicoccin (FC) stimulated the increase in fresh weight but not the increase in the RNA and DNA levels; in this condition, the inhibitor level decreased and the increase in Cam level was higher than in the control. In the presence of both ABA and FC, there was an increase in fresh weight not accompanied by an increase in DNA and RNA levels; Cam increased and, on a fresh weight basis, reached the value of the control. These results indicate that the Ca(2+)-Cam system was activated in early germination of radish seeds by an increase in Cam and a decrease in the inhibitor levels, that FC, probably through the activation of membrane functions, increased Cam level, and that the ABA inhibition on germination was not mediated by the Ca(2+)-Cam system.

Journal Article↗
PubMed1989

A beta-Galactosidase from Radish (Raphanus sativus L.) Seeds.

A basic beta-galactosidase (beta-Galase) has been purified 281-fold from imbibed radish (Raphanus sativus L.) seeds by conventional purification procedures. The purified enzyme is an electrophoretically homogeneous protein consisting of a single polypeptide with an apparent molecular mass of 45 kilodaltons and pl values of 8.6 to 8.8. The enzyme was maximally active at pH 4.0 on p-nitrophenyl beta-d-galactoside and beta-1,3-linked galactobiose. The enzyme activity was inhibited strongly by Hg(2+) and 4-chloromercuribenzoate. d-Galactono-(1-->4)-lactone and d-galactal acted as potent competitive inhibitors. Using galactooligosaccharides differing in the types of linkage as the substrates, it was demonstrated that radish seed beta-Galase specifically split off beta-1,3- and beta-1,6-linked d-galactosyl residues from the nonreducing ends, and their rates of hydrolysis increased with increasing chain lengths. Radish seed and leaf arabino-3,6-galactan-proteins were resistant to the beta-galase alone but could be partially degraded by the enzyme after the treatment with a fungal alpha-l-arabinofuranosidase leaving some oligosaccharides consisting of d-galactose, uronic acid, l-arabinose, and other minor sugar components besides d-galactose as the main product.

Journal Article↗
PubMed1990

A Two-Dimensional Vibrating Probe Study of Currents around Lateral Roots of Raphanus sativus Developing in Culture.

A computer-assisted, two-dimensional vibrating probe was used to study the ionic currents around developing lateral roots of Raphanus sativus in vitro. This system allowed us to superimpose current vectors on the video image of the roots. In a young lateral root, current entered the cap, meristematic, and elongation zones and exited the primary root surface close to the base of the lateral root. As the lateral root grew, current began to exit from its basal (cell maturation zone) end. The densities of currents entering the apical portion of the faster-growing lateral roots in a medium lacking indole 3-acetic acid were about twice as large as those entering the apical region of the slower-growing lateral roots in indole 3-acetic acid-supplemented medium.

Journal Article↗
PubMed1995

Molecular and cytological characterization of a highly repeated DNA sequence in Raphanus sativus.

A highly repeated DNA sequence with a repeat unit of ca. 180 bp was found in genomic DNA HindIII-digests of Raphanus sativus. The repeating units of six isolated, independent clones were sequenced. These units have 177 or 178 bp, are 36% G+C in their DNA base composition, and show 90% sequence homology. The copy number of this 180-bp repeat unit is about 0.5 x 10(6) per diploid genome. In situ hybridization analysis with the repeating units as the probe and C-banding analysis indicated that the repeated DNA sequence of R. sativus is closely associated with the major C-heterochromatins in the proximal regions of all 18 chromosomes at mitotic metaphase.

Base Composition↗
PubMed1991

Malate content of picoliter samples of Raphanus sativus cytoplasm.

Malate, which plays many essential roles in plant metabolism, is a potent in vitro inhibitor of the cytosolic enzyme phosphoenolpyruvate carboxylase (PEPC). Because PEPC activity leads to malate biosynthesis, malate is assumed to attenuate its own synthesis in situ. To test this hypothesis, we measured directly the malate content of picoliter samples of Raphanus root-hair cytoplasm using quantitative histochemical techniques. We also obtained an estimate for malate accumulation in these cells. These values were compared with the PEPC activity of individual root hairs (less than 2 ng). The results indicate that high cytoplasmic malate concentration does not severely inhibit PEPC in situ. We suggest that the focus for studies on the regulation of organic anion accumulation be on the interactive effects of malate and other PEPC effectors.

Cytoplasm↗
PubMed1976

An ultrastructural and radioautographic study of the chromocentric interphase nucleus in plant meristematic cells (Raphanus sativus).

In Raphanus sativus, the mitotic chromosomes are quite short and, on reaching the cell poles, soon undergo extensive unravelling. By late telophase and early interphase, only a few chromosome segments, believed to correspond to the centromeric regions, are still visible in the form of chromocentres closely associated with the nuclear envelope. Although interphase nuclei show little internal structural differentiation, high-resolution radioautography has permitted us to establish which of them have reached the early, mid and late S periods. In early S nuclei, only the nucleolus and the euchromatin which pervades the nuclear cavity become labelled. By the mid S-period, the diffuse chromatin and nucleolus incorporate less thymidine and DNA synthesis is initiated within the peripheral chromocentres. Subsequently, the radioautographic grains become restricted to the chromocentres. The finding that certain late S nuclei exhibit loosely organized chromocentres strongly suggests that these heterochromatic chromosome segments undergo important conformational modifications during DNA replication. Finally, the presence of radioautographic grains over the lacunar regions of the nucleolus in early and mid S nuclei demonstrates that intranucleolar DNA replicates during the earlier portion of the S-period.

Autoradiography↗
PubMed2001

Allozyme diversity and population structure of Japanese and Korean populations of wild radish, Raphanus sativus var. hortensis f. raphanistroides (Brassicaceae).

Raphanus satiuus var. hortensis f. raphanistroides (wild radish: Brassicaceae) is an insect-pollinated wild plant that grows mainly on beaches in East Asia. Starch gel electrophoresis was used to investigate the allozyme diversity and genetic structure of 25 Japanese and 9 Korean populations of this plant. Although the Korean populations were small, isolated, and patchily distributed, they maintained a high level of genetic diversity; the average percentage of polymorphic loci was 63.1%, the mean number of alleles per locus was 2.27, and the average heterozygosity was 0.278. The corresponding estimates for these parameters in the Japanese populations were 53.3%, 2.26, and 0.278. These estimates are considerably higher than those from species with similar life history and ecological characteristics, but they are lower than those from R. raphanistrum, the wild radish that grows in Europe and the U.S.A. The combination of an insect-pollinated, outcrossing breeding system, large population sizes, gene flow from cultivated radish population, and a propensity for high fecundity may explain the high level of genetic diversity within wild populations.

Alleles↗
PubMed1985

Effects of feeding wheat contaminated with wild radish (Raphanus raphanistrum) to growing pigs.

In each of two growth trials, weanling pigs were individually housed and fed to study the effect of various amounts of wild radish (Raphanus raphanistrum) seed pods in the diet on performance of growing swine. For trial 1, 20 pigs averaging 19.6 kg live weight were divided into four comparable groups of five pigs each based on sex and initial weight and were assigned to fortified wheat-soybean meal diets with: (1) 0%, (2) 1.20%, (3) 3.25%, or (4) 4.95%, by weight, of wild radish seed pods. For trial 2, 15 pigs averaging 20.6 kg live weight were divided into three comparable groups based on sex and initial weight and assigned to fortified wheat-soybean meal diets with: (5) 0%, (6) 7.5%, or (7) 15% of wild radish seed pods. Average daily gains of pigs during the 60 d of trial 1 on all diets were similar (P greater than .05). However, wild radish contamination of the diet had a significant linear effect on feed conversion because the amount of feed required per unit of gain increased (P less than .01) in the amount of feed required to produce a unit of gain. There were no gross symptoms of toxicity in any pigs during the course of either trial or at maturity. Eighteen crossbred barrows averaging 24.8 kg were divided into three comparable groups of six pigs each and assigned to each of the three diets (5, 6 and 7) fed to growing pigs in trial 2, for a digestion and nitrogen balance trial.(ABSTRACT TRUNCATED AT 250 WORDS)

Animal Feed↗
PubMed1999

Inhibition of growth of Escherichia coli O157:H7 in fresh radish (Raphanus sativus L.) sprout production by calcinated calcium.

The inhibitory effect of calcinated calcium on the growth of Escherichia coli O157:H7 during fresh radish (Raphanus sativus L.) sprout production was studied. It was revealed that the addition of 0.4% (wt/vol) calcinated calcium into radish sprouting medium which was artificially contaminated with E. coli O157:H7 (3.0 to 3.2 log CFU/ml) completely inhibited the growth or inactivated the microorganism. When radish seed extract was used instead of radish sprout production, the same extent of growth inhibition or inactivation was observed with much lower amounts (0.07%) of calcinated calcium under similar experimental conditions. The findings suggested that calcinated calcium may be useful to control E. coli O157:H7 contamination during the production of radish sprouts.

Brassicaceae↗
PubMed1976

[Virus infestation of garden radish (Raphanus sativus L. var. sativus) (author's transl)].

In July 1974 a mosaic disease was stated in seed plants of garden radish near Aschersleben which spread more and more within some weeks and eventually infected over 40 per cent of the stand. Investigations in comparison with healthy seed plants demonstrated the following average losses induced by the disease: number of pods 51 per cent, seed weight 67 per cent, number of seeds per plant 53 per cent, weight of 1,000 seeds 33 per cent. Nearly unaffected were: number of seeds per pod, percentage of seed germination, and percentage of emerged seedlings. However, the seedlings were smaller when they derived from diseased mother plants. From diseased seed plants cauliflower mosaic virus was isolated. Seed transmission of this virus could not be stated with several hundreds of seedlings deriving from diseased garden radish. Back transmissions were successful with sap and with aphids. Only one strain which was cultivated in greenhouse for a long time gave no positive results. Three isolates of cabbage black ring virus from different plant species and the four beetle-transmissible viruses, namely turnip yellow mosaic, turnip crinkle, turnip rosette, and radish mosaic viruses induced no symptoms on garden radish seedlings. However, Erysimum latent virus, also belonging to the beetle-transmissible viruses, sometimes caused symptoms in garden radish. For the first time the investigations demonstrated in Europe, but probably also for the other continents, a spontaneous virus infection of Raphanus sativus var. sativus.

Animals↗