Molecular characterization of a new type of cytoplasmic male sterile sugar beet

A line (named Cl) of cytoplasmic sterility of sugar beet whose cytoplasm derived from Betacicla Turkey was obtained by interspecific hybrid. Its cytoplasm and a spontaneous male sterile cytoplasm from wild beet Beta maritima (named M) were compared with that of Owen's sterile line (S-cms) and a common maintainer of them named N was used as control. RFLP and RAPD methods were mainly used in our experiments. The restriction fragment patterns of mtDNAs were found to be likely but for a few of specific low-lighted electrophoresis bands in Cl. The results of Southern hybridization of six heterogeneous mitochondrial genes as probes to digests of mtDNAs by six restriction enzymes showed to be analogous between S and M lines. But the Cl mtDNA was sorted out by hybridization of atpA probe. Difference of low-molecular-weight mitochondrial DNAs was found among the three sterile lines. Three RNA molecules weighing about 4.2kb stably existed in Cl mitochondria. Our results of RAPD also supported that the Cl cytoplas     

Direct and indirect effects of Cd2+ on photosynthesis in sugar beet (Beta vulgaris)

Sugar-beet plants ( Beta vulgaris L. cv. Monohill) were cultivated for 4 weeks in a complete nutrient solution. Indirect effects of cadmium were studied by adding 5, 10 or 20 μ M CdCl₂ to the culture medium while direct effects were determined by adding 1, 5, 20, 50 or 2 000 μ M CdCl₂ to the assay media. The photosynthetic properties were characterized by measurement of CO₂ fixation in intact plants, fluorescence emission by intact leaves and isolated chloroplasts, photosystem (PS) I and PSII mediated electron transport of isolated chloroplasts, and CO₂-dependent O₂ evolution by protoplasts. When directly applied to isolated leaves, protoplasts and chloroplasts. Cd²⁺ impeded CO₂ fixation without affecting the rates of electron transport of PSI or PSII or the rate of dark respiration. When Cd²⁺ was applied through the culture medium the capacity for, and the maximal quantum yield of CO₂ assimilation by intact plants both decreased. This was associated with: (1) decreased total as well as effective chlorophyll content (PSII antennae size), (2) decreased coupling of electron transport in isolated chloroplasts, (3) perturbed carbon reduction cycle as indicated by fluorescence measurements. Also, protoplasts isolated from leaves of Cd²⁺-cultivated plants showed an increased rate of dark respiration.     

An extended map of the sugar beet genome containing RFLP and RAPD loci

An updated map of sugar beet (Beta vulgaris L. ssp. vulgaris var altissima Doell) is presented. In this genetic map we have combined 248 RFLP and 50 RAPD loci. Including the loci for rhizomania resistance Rr1, hypocotyl colour R and the locus controlling the monogerm character M, 301 loci have now been mapped to the nine linkage groups covering 815 cM. In addition, the karyotype of some of the Beta vulgaris chromosomes has been correlated with existing RFLP and RAPD linkage maps.     

Plant regeneration from protoplasts of sugar beet (Beta vulgaris)

Mesophyll protoplasts from two of five sugar beet lines tested were regenerated into plants. Mesophyll protoplasts of all lines showed high plating efficiencies up to 4.0% developed hard compact callus, and two of the lines also developed white, soft and friable callus consisting of starch grain-containing cells. Whereas the compact callus never regenerated into plants, the white friable ones frequently developed globular structures, which became green in the light and formed adventitious shoots after cytokinin (BAP or thidiazuron) treatment. Genetic analysis by PCR-fingerprinting and flow cytometry showed uniformity and unchanged ploidy levels in 15 independently regenerated plantlets in line NF. but altered ploidy level (from diploid to triploid) in a regenerated plantlet of clone VRB.     

Mobilisation of storage reserves during germination and early seedling growth of sugar beet

The principal storage reserve of sugar beet seeds is starch, which is localised in the perisperm. Additional storage reserves include the seed proteins, albumins, globulins and glutclins, which are exclusively located in the embryo. Soluble sugars are also detectable in all the organs of the mature seed. The time-course of reserve mobilisation in the different organs of the sugar beet ( Beta vulgaris L. cv. Regina) seed during germination and early seedling growth is documented, with particular reference to changes in (a) activities of hydrolases: a-amylase, β-amylase, and α-glucosidases; (b) levels of carbohydrates and (c) proteins. Amylase activities increase substantially in both cotyledons, as well as the perisperm, whereas the increase in α-glucosidase activities is largely confined to the perisperm.     

Monitoring gene flow from transgenic sugar beet using cytoplasmic male-sterile bait plants

One of the most discussed environmental effects associated with the use of transgenic plants is the flow of genes to plants in the environment. The flow of genes may occur through pollen since it is the reproductive system that is designed for gene movement. Pollen-mediated gene escape is hard to control in mating plants. Pollen from a wind pollinator can move over distances of more than 1000 m. To investigate the efficiency of transgenic pollen movement under realistic environmental conditions, the use of bait plants might be an effective tool. In this study, cytoplasmic male-sterile (CMS) sugar beets were tested with regard to their potential for monitoring transgene flow. As the pollen source, transgenic sugar beets were used that express recombinant DNA encoding viral (beet necrotic yellow vein virus) resistance, and antibiotic (kanamycin) and herbicide (glufosinate) tolerance genes. In a field trial, the effectiveness of a hemp (Cannabis sativa) stripe containment strategy was tested by measuring the frequency of pollinated CMS bait plants placed at different distances and directions from a transgenic pollen source. The results demonstrated the ineffectiveness of the containment strategy. Physiological and molecular tests confirmed the escape and production of transgenic offspring more than 200 m behind the hemp containment. Since absolute containment is unlikely to be effective, the CMS-bait plant detection system is a useful tool for other monitoring purposes.     

Cadmium effects on leaf transpiration of sugar beet (Beta vulgaris)

Seedlings of sugar beet ( Beta vulgaris L. cv Monohill) were cultivated for 4 weeks in nutrient solution containing different concentrations of CdCl₂ (0 to 10 μ M ). The effects of Cd on appearance and function of stomata and leaf cuticle were investigated by water loss measurements and microscopy. The leaf transpiration rate increased with increasing Cd concentrations while the sum total of stomatal aperture area per unit leaf area decreased. Already at low Cd levels. an increase of defective and undeveloped stomata was found in Cd treated plants. These stomata are closed or have small apertures and probably lack a functional closing mechanism. The number of intact stomata per unit leaf area was lower in leaves of Cd treated plants than in controls, and Cd induced closure of intact stomata. The total number of stomata per leaf area slightly increases with increasing Cd concentration. as does the percentage of small stomata. Furthermore. specific leaf area increased, while the density of leaf structure was decreased by Cd. From this observation we conclude that the increase in transpiration rate caused by Cd is primarily due to effects on the permeability of the leaf cuticle to water.     

Carbohydrate starvation is a major determinant of the loss of greening capacity in cotyledons of dark-grown sugar beet seedlings

The transition of cotyledons from heterottophy to autotrophy is a critical step of seedling establishment. We have studied the greening capacity of sugar beet ( Beta vulgaris L. cv. Véga) cotyledons in relation to carbohydrate and energy metabolism during dark growth. During early growth, sugar beet cotyledons behaved mainly as a lipid-mobilizing and gluconeogenic tissue providing substrates to the seedling. Reserve mobilization was followed by a maximum of the adenine nucleotide pool on day 6 in strict correlation with the maximum of greening capacity. This was immediately followed by the onset of a typical situation of carbohydrate starvation characterized by substrate limitation of respiration, a decrease in the adenine nucleotide pool and, as shown by the respiratory quotient and the loss of proteins, a probable utilization of cellular proteins and lipids to sustain respiration. The conversion from etioplast to chloroplast, as determined by the rate of chlorophyll synthesis, was less and less efficient as carbohydrate starvation continued, finally leading to incomplete and heterogeneous greening on day 12. The relationship of the loss of greening capacity with carbohydrate starvation is discussed.     

Calcium reduces toxicity of aminoglycoside antibiotics in sugar beet explants in vitro

Aminoglycoside antibiotics are frequently used for the selection of transgenic plant cells. However, for a number of species aminoglycoside selection is inefficient. The objective of the present study was to elucidate factors affecting the phytoloxic effects of aminoglycoside antibiotics. Using non-transgenic sugar beet cotyledonary explants the interaction between three aminoglycoside antibiotics, kanamycin, neomycin and hygromycin. and Ca²⁺ was studied by monitoring the effects on growth and shoot formation. The phytotoxic effects of the aminoglycoside antibiotics were strongly dependent on the calcium concentration in the growth media. At comparable levels of the antibiotics (kanamycin 170 μ M , neomycin 220 μ M , hygromycin 9.5 μ M , an elevation of the calcium concentration from 1 to 10 m M resulted in growth increases of approximately 3-, 2.5- and 8-fold, respectively, and shoot formation was enhanced 1.5-, 2-and 6-fold, respectively. At lower concentrations of the antibiotics, the toxic effect was nearly abolished by increasing the calcium concentration. Additional magnesium, sodium and ammonium did not affect the phytotoxic effects of the aminoglycoside antibiotics. Moreover, the phytotoxic effects of the herbicides glyphosate and phosphinothricin were not decreased by additional calcium. These data suggest the existence of a specific interaction between calcium and aminoglycoside anfibiotics in plants. The implications of these results for the use of aminoglycosides as selective agents in plant transformation are discussed.     

An analysis of genetic variation in sugar beet (Beta vulgaris L. using an augmented biparental (ABIP) mating design

An augmented biparental (ABIP) mating design was used to investigate the quantitative variation, particularly the dominance variation, for morphological and chemical characters in sugar beet. Diploid O-type plants were both crossed and selfed and the progeny were grown in a single-plant randomised field trial. A comparison of the two kinds of family provided tests for both dominance variation and directional dominance effects. Estimates of the narrow heritability were also obtained for each character. Germination problems reduced the size of the final analysis but evidence was obtained of dominance variation and positive directional dominance effects for leaf length, root weight and potassium concentration and, to a lesser extent, sugar concentration. Genetic control of sodium and alpha-amino nitrogen concentrations appeared to be mostly additive. Hybrid varieties of sugar beet should exploit these directional dominance effects and the more closely varieties approach true F, hybrids the more they will capitalise on these advantages. It is possible that other factors such as epistasis, contamination, competition and seed effects may cause us to under- or overestimate the importance of dominance.     

Protein synthesis in mitochondria purified from roots, leaves and flowers of sugar beet

Highly purified, intact and functional mitochondria were isolated from roots and leaves of a number of fertile and male-sterile lines of sugar beet ( Beta vulgaris L.). Intact and functional mitochondria were successfully isolated from the flowers of fertile plants, but not from the flowers of male-sterile plants. Several alternative methods for the homogenization of male-sterile flowers were tried. Their failure suggests that the mitochondria from male-sterile flowers are more sensitive to mechanical damage than mitochondria from fertile, or other organs of male-sterile, plants.
In organello protein synthesis was optimized with respect to the total concentration of amino acids, the concentration of [³⁵S]-methionine, pH and respiratory substrate. Inhibitor experiments showed that the mitochondrial preparations contained mitochondrial translational activity only. With the exception of one band, no processing or proteolytic breakdown in either root or leaf mitochondrial protein synthesis products could be detected in pulse-chase experiments. Submitochondrial fractionation experiments showed the presence of two soluble polypeptides, whereas all other polypeptides were membrane bound.
The polypeptide patterns of root, leaf and flower mitochondria were very similar with the exception of 4 polypeptides involved in glycine oxidation. These 4 polypeptides were present in large amounts in leaf mitochondria and just detectable in flower mitochondria. The patterns of polypeptides syntesized in mitochondria isolated from roots, leaves and flowers also showed a number of organ-specific differences. Six qualitative and 6 quantitative differences were found between mitochondria isolated from these three organs. No unique polypeptides were found to be synthesized either by flower mitochondria or by mitochondria from roots and leaves of male-sterile plants compared to their male-fertile counterparts.     

Selection in vitro for UV-tolerant sugar beet (Beta vulgaris) somaclones

With a reduced stratospheric ozone concentration, the generation of UV-tolerant plants may be of particular importance. Among different crop plants there is large variation in sensitivity to UV-B radiation. This study was undertaken to investigate the possibilities of using somaclonal variation and selection in vitro for improving UV-B tolerance in sugar beet (Beta vulgaris L.). Sugar beet callus was exposed to UV radiation (280–320 nm, 0.863–5.28 kJ m^-2 day^-1, unweighted) and resultant shoots were selected from surviving cells. After establishment of the plants, they were grown under either visible radiation (114 μmol m^-2 s^-1 PAR) or with the addition of UV radiation (6.3 kJ m^-2 day^-1 biologically effective UV-B). Screening of regenerants in vivo for tolerance to UV radiation was undertaken 10 months after termination of the UV selection pressure. Screening was done visually and by using a number of physiological parameters, including chlorophyll fluorescence induction, ultraweak luminescence, pigment analysis and total content of UV-screening pigments. A clear difference between the unselected and the UV-selected somaclones was observed when visually studying the UV damage and other leaf injury. The observations were supported by the ultraweak luminescence measurements. Unselected plants showed significantly greater damage when subjected to subsequent UV radiation as compared to the selected plants. The clones subjected to UV selection pressure displayed a significantly higher concentration of UV-screening pigments under subsequent UV radiation. The unselected plants under subsequent UV treatment showed a lower carotenoid concentration when compared to selected plants. However, no significant difference between treatments was found for chlorophyll a/b, or F/F_max, a measure of photosynthetic quantum yield.     

A linkage map of sugar beet (Beta vulgaris L.)

Summary We have established a first linkage map for beets based on RFLP, isozyme and morphological markers. The population studied consisted of 96 F₂ individuals derived from an intraspecific cross. As was expected for outbreeding species, a relatively high degree of polymorphism was found within sugar beet; 47% of the DNA markers were polymorphic for the chosen population. The map consists of 115 independent chromosomal loci designated by 108 genomic DNA probes, 6 isozyme and one morphological marker. The loci cover 789 cM with an average spacing of 6.9 cM. They are dispersed over nine linkage groups corresponding to the haploid chromosome number of Beta species. Eighteen markers (15.4%) showed distorted segregation which, in most instances, can be explained by gametic selection of linked lethal loci. The application of the linkage map in sugar beet breeding is discussed.     

Molecular markers applied to plant tissue culture

Summary The last decade has witnessed successful applications of plant tissue culture techniques in several crops. During that same period, studies in plant molecular genetics have also grown exponentially. Molecular markers (isozymes, RFLPs, and PCR-based markers such as RAPDs) are now used to study many of the current limitations of tissue culture. They have been used to investigate mechanisms that underlie somaclonal variation in the nuclear, mitochondrial, and chloroplast genomes. One recurrent problem with several tissue culture systems has been the difficulty of determining the origin of regenerants. Molecular markers represent powerful tools to determine precisely the origin of plants derived from microspore or anther culture, protoplast fusion, and other tissue culture studies where this information is important. With improvements in tissue culture techniques, populations of doubled haploid lines have been produced in several major crop species. Doubled haploid populations have proven useful in the production of molecular maps and in tagging important agronomic traits. This review describes the use of molecular markers to address fundamental and practical questions of plant tissue culture, and discusses the potential of improvements in molecular techniques and new molecular markers such as SCAR and STS along with high-resolution mapping strategies.     

A comparison between ethylene production, ACC and mACC contents, and hydroperoxide level in normal and habituated sugar beet calli

In contrast to normal hormone, requiring sugar beet callus, habituated auxin- and cytokinin-independent callus of the same plant produces very little ethylene, contains less 1-aminocyclopropane-1-carboxylic acid (ACC) and malonyl-ACC (mACC), has a low capacity to convert ACC into ethylene and has lower levels of hydroperoxides. The low ethylene production is apparently controlled by the rate of ACC synthesis and conversion to mACC, as well as by the activity of the ethylene forming enzyme. The interactions between ethylene and polyamine metabolism are discussed, as well as the possible causal relation between the low level of ethylene and the low degree of differentiation of the habituated cells.     

Transcriptome analysis of sugar beet root maggot (Tetanops myopaeformis) genes modulated by the Beta vulgaris host

Sugar beet root maggot (SBRM, Tetanops myopaeformis von Röder) is a major but poorly understood insect pest of sugar beet (Beta vulgaris L.). The molecular mechanisms underlying plant defense responses are well documented, however, little information is available about complementary mechanisms for insect adaptive responses to overcome host resistance. To date, no studies have been published on SBRM gene expression profiling. Suppressive subtractive hybridization (SSH) generated more than 300 SBRM ESTs differentially expressed in the interaction of the pest with a moderately resistant (F1016) and a susceptible (F1010) sugar beet line. Blast2GO v. 3.2 search indicated that over 40% of the differentially expressed genes had known functions, primarily driven by fruit fly D. melanogaster genes. Expression patterns of 18 selected EST clones were confirmed by RT‐PCR analysis. Gene Ontology (GO) analysis predicted a dominance of metabolic and catalytic genes involved in the interaction of SBRM with its host. SBRM genes functioning during development, regulation, cellular process, signaling and under stress conditions were annotated. SBRM genes that were common or unique in response to resistant or susceptible interactions with the host were identified and their possible roles in insect responses to the host are discussed.     

The design and analysis of breeders' trials of sugar beet (Beta vulgaris) using two-dimensional blocking structures

A method for generating two-dimensional blocking designs to fit any shape and size of experimental area is presented. The method takes an alpha design appropriate to the experimental area and imposes additional blocking perpendicular to the alpha blocks. Improvements to the design are then made by repositioning entries within the alpha blocks. Although this method of construction is less sophisticated than for other two-dimensional designs, the designs are particularly suited to large scale breeders trials where no alternative two-dimensional design may exist. The designs have been used for 3 yr in a sugar beet breeding programme, and have given improvements in efficiency over one-dimensional alpha designs.     

甜菜纤维的制备及其性质

甜菜纤维的制备及其性质樊志和周人纲王占武李晓芝韩炜（河北省农林科学院农业物理生理生化研究所,石家庄０５００５１）Ａｐｒａｃｔｉｃａｌｐｒｅｐａｒａｔｉｏｎａｎｄｃｈａｒａｃｔｅｒｉｚａｔｉｏｎｏｆｄｉｅｔａｒｙｆｉｂｅｒｆｒｏｍｓｕｇａｒ┐ｂｅｅｔｐ...     

Polyamine metabolism and ethylene biosynthesis in normal and habituated sugar beet callus

The optimal assay conditions and the trend with time in culture (28 days) of arginine decarboxylase (ADE; EC 4.1.1.19), omithine decarboxylase (ODC; EC 4.1.1.17) and diamine oxidase (DAO; EC 1.4.3.6) activities in habituated (H) and normal (N) auxin- and cytokinin-requiring sugar beet callus were compared. Although the response to variations in buffer pH and EDTA and pyridoxal phosphate (PLP) concentrations varied for ADC and ODC activities between the two callus types, pH 8.3, 50 μ M PLP and 5 m M EDTA were generally optimal or near-optimal for both H and N callus. In most cases the addition of ornithine or arginine in the ADC and ODC assays, respectively, given to block the interconversion between the two substrates, resulted in lower ¹⁴CO₂ recovery. DAO activity was very differently affected in H and N callus by the presence of polyvinylpyrrolidone in the extration buffer. However, in both cases, this activity increased with time in culure. ADC activity was always predominant in both cell lines and always higher in N callus. In the latter, ADC activity rose sharply between days 14 and 21 and then leveled off while in H callus it incresed steadily from day 14 onwards. ODC activity was also higher in N callus and peaked sharply on day 21 while in H callus it was not detectable in the second half of the culture period. In both cell lines this activity was low or nil on day 28. 3,4-[¹⁴C]-methionine incorporation into ethylene and polyamines was also compared in N and H callus. In the latter, ethylene synthesis was lower and [¹⁴C]-spermidine formation higher than in N callus. This is in accord with the significantly higher spermidine titres found in H callus.