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.     

Genetic approaches to sustainable pest management in sugar beet (Beta vulgaris)

Sugar beet (Beta vulgaris) is an important arable crop, traditionally used for sugar extraction, but more recently, for biofuel production. A wide range of pests, including beet cyst nematode (Heterodera schachtii), root‐knot nematodes (Meloidogyne spp.), green peach aphids (Myzus persicae) and beet root maggot (Tetanops myopaeformis), infest the roots or leaves of sugar beet, which leads to yield loss directly or through transmission of beet pathogens such as viruses. Conventional pest control approaches based on chemical application have led to high economic costs. Development of pest‐resistant sugar beet varieties could play an important role towards sustainable crop production while minimising environmental impact. Intensive Beta germplasm screening has been fruitful, and genetic lines resistant to nematodes, aphids and root maggot have been identified and integrated into sugar beet breeding programmes. A small number of genes responding to pest attack have been cloned from sugar beet and wild Beta species. This trend will continue towards a detailed understanding of the molecular mechanism of insect–host plant interactions and host resistance. Molecular biotechnological techniques have shown promise in developing transgenic pest resistance varieties at an accelerated speed with high accuracy. The use of transgenic technology is discussed with regard to biodiversity and food safety.     

Intraspecific diversity of sugar beet (Beta vulgaris) mitochondrial DNA

Summary Mitochondrial (mt) DNA, isolated from different sugar beet populations, was analyzed using BamHI and EcoRI restriction enzymes. It was shown that plants possessing the new mtDNA types are revealed among O-type fertilizers quite frequently. Among cytoplasmic male sterile (cms) plants, which evolved during cultivation of O-type fertilizers, plants with altered mt genome were found.     

Genetic analysis of bolting after winter in sugar beet (Beta vulgaris L.)

Key message

This study reveals for the first time a major QTL for post-winter bolting resistance in sugar beet ( Beta vulgaris L.). The knowledge of this QTL is a major contribution towards the development of a winter sugar beet with controlled bolting behavior.

Abstract

In cool temperate climates, sugar beets are currently grown as a spring crop. They are sown in spring and harvested in autumn. Growing sugar beet as a winter crop with an extended vegetation period fails due to bolting after winter. Bolting after winter might be controlled by accumulating genes for post-winter bolting resistance. Previously, we had observed in field experiments a low post-winter bolting rate of 0.5 for sugar beet accession BETA 1773. This accession was crossed with a biennial sugar beet with regular bolting behavior to develop a F₃ mapping population. The population was grown in the greenhouse, exposed to artificial cold treatment for 16 weeks and transplanted to the field. Bolting was recorded twice a week from May until October. Post-winter bolting behavior was assessed by two different factors, bolting delay (determined as days to bolt after cold treatment) and post-winter bolting resistance (bolting rate after winter). For days to bolt, means of F₃ families ranged from 25 to 164 days while for bolting rate F₃ families ranged from 0 to 1. For each factor one QTL explaining about 65 % of the phenotypic variation was mapped to the same region on linkage group 9 with a partially recessive allele increasing bolting delay and post-winter bolting resistance. The results are discussed in relation to the potential use of marker-assisted breeding of winter sugar beets with controlled bolting.     

Intraspecific variation of Myzus persicae on sugar beet (Beta vulgaris)

Differences in inherited resistance among seven sugar-beet stocks had similar effects on Myzus persicae clones representing the range of variation in aphid response to resistant and susceptible sugar beet observed in fifty-eight clones collected between 1969 and 1971. Three sugar-beet stocks were consistently resistant. Statistically significant interactions between beet stocks and aphid clones did not indicate the existence of biotypes with specific abilities to overcome resistance. M. persicae clones differed in their vigour of colonizing sugar beet, irrespective of the differences between beet stocks. The readiness of adult aphids to settle determined the size of aphid population produced and included a component related to the response of the aphid clone to sugar beet as a host, and a component related to the resistance ranking of the beet stock. Breeding sugar beet with resistance to aphids will be simplified, as the results indicate that, at present, differences between aphid biotypes need not be considered a problem.     

Molecular profiling of microbial communities associated with seeds of Beta vulgaris subsp. Vulgaris (sugar beet)

The composition of microbial communities on and within seeds may effect their storage and field performance, whether they are indigenous or applied as biocontrol agents. In this study, we have explored the usefulness of profiling small subunit ribosomal (SSR) gene fragments for studying the microflora associated with seeds. DNA was amplified by the polymerase chain reaction (PCR) and the amplicons separated using denaturing gradient gel electrophoresis (DGGE). Primers targeting eukaryotic SSRs were used to investigate fungal communities, and primers targeting bacterial SSRs were employed to study the eubacterial microflora. As a case study, we attempted to profile the fungi and bacteria associated with seeds of Beta vulgaris (sugar beet) to permit an insight into the varying field performance of several well-characterised commercial seed lots. Serious interference with the microbial signals was observed from the plant's own nuclear 18S rRNA genes and chloroplast 16S rRNA genes using standard PCR conditions and DNA extracted from whole seeds as template. Hot-start and touchdown PCR made no appreciable improvement to these signals. Seed imbibition and dissection into operculum and fruit wall and true seed prior to DNA extraction improved signal recovery in the fruit fraction. With primer modification, bacteria and fungi were detected in an excess of plant DNA of 100:1 and 10:1, respectively. With this method, microbial communities on seeds could be profiled, however, it is likely that targeted depletion of plant rDNA targets will be a necessary extra step before this approach can be used to screen seeds routinely.     

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.     

Flavin excretion from roots of iron-deficient sugar beet (Beta vulgaris L.)

The characteristics of flavin excretion from iron-deficient sugar-beet roots have been studied. Roots from iron-deficient sugar beet excreted flavins when plants were allowed to decrease the pH of the nutrient solution, but not when plants were grown in nutrient solutions buffered at high pH. As shown by reversed-phase high-performance liquid chromatography, the two major flavins whose excretion was induced by iron deficiency were different from riboflavin, FMN and FAD. These flavins have been identified as riboflavin 3′-sulfate and riboflavin 5′-sulfate by electrospray-mass spectrometry, inductively coupled plasma emission spectroscopy, infrared spectrometry and¹H-nuclear magnetic resonance. We have characterized the time courses of accumulation of the different flavins in the nutrient solution and considered several possible roles for flavin excretion in iron acquisition.     

Genetic localization of four genes for nematode (Heterodera schachtii Schm.) resistance in sugar beet (Beta vulgaris L.)

Sugar beet (Beta vulgaris L.) is highly susceptible to the beet cyst nematode (Heterodera schachtii Schm.). Three resistance genes originating from the wild beets B. procumbens (Hs1 ^pro-1) and B. webbiana (Hs1 ^web-1, Hs2 ^web-7) have been transferred to sugar beet via species hybridization. We describe the genetic localization of the nematode resistance genes in four different sugar beet lines using segregating F₂ populations and RFLP markers from our current sugar beet linkage map. The mapping studies yielded a surprising result. Although the four parental lines carrying the wild beet translocations were not related to each other, the four genes mapped to the same locus in sugar beet independent of the original translocation event. Close linkage (0–4.6 cM) was found with marker loci at one end of linkage group IV. In two populations, RFLP loci showed segregation distortion due to gametic selection. For the first time, the non-randomness of the translocation process promoting gene transfer from the wild beet to the sugar beet is demonstrated. The data suggest that the resistance genes were incorporated into the sugar beet chromosomes by non-allelic homologous recombination. The finding that the different resistance genes are allelic will have major implications on future attempts to breed sugar beet combining the different resistance genes.     

dsRNA-mediated resistance to Beet Necrotic Yellow Vein Virus infections in sugar beet (Beta vulgaris L. ssp. vulgaris)

Rhizomania, one of the most devastating diseases in sugar beet, is caused by Beet Necrotic Yellow Vein Virus (BNYVV) belonging to the genus Benyvirus. Use of sugar beet varieties with resistance to BNYVV is generally considered as the only way to maintain a profitable yield on rhizomania-infested fields. As an alternative to natural resistance, we explored the transgenic expression of viral dsRNA for engineering resistance to rhizomania. Transgenic plants expressing an inverted repeat of a 0.4 kb fragment derived from the BNYVV replicase gene displayed high levels of resistance against different genetic strains of BNYVV when inoculated using the natural vector, Polymyxa betae. The resistance was maintained under high infection pressures and over prolonged growing periods in the greenhouse as well as in the field. Resistant plants accumulated extremely low amounts of transgene mRNA and high amounts of the corresponding siRNA in the roots, illustrative of RNA silencing as the underlying mechanism. The transgenic resistance compared very favourably to natural sources of resistance to rhizomania and thus offers an attractive alternative for breeding resistant sugar beet varieties.     

Genetic and chromosomal localization of the 5S rDNA locus in sugar beet (Beta vulgaris L.).

A digoxigenin-labelled 5S rDNA probe containing the 5S rRNA gene and the adjacent intergenic spacer was used for in situ hybridization to metaphase and interphase chromosomes of a trisomic stock from sugar beet (Beta vulgaris L.). Three chromosomes of primary trisomic line IV (T. Butterfass. Z. Bot. 52: 46-77. 1964) revealed signals close to the centromeres. Polymorphisms of 5S rDNA repeats in a segregating population were used to map genetically the 5S rRNA genes within a cluster of markers in linkage group II of sugar beet. The concentration of genetic markers around the centromere presumably reflects the suppressed recombination frequency in centromeric regions. The correlation of physical and genetic data allowed the assignment of a linkage group to sugar beet chromosome IV according to line IV of the primary trisomics.     

Predicting the seedling emergence time of sugar beet (Beta vulgaris) using beta models

Soil temperature, texture, water content and sowing depth are effective factors on the estimation of emergence time. This research aimed to test the Beta model for its adequacy in predicting the time of emergence for sugar beet. The Beta growth model as a phenological model have been used for evaluating the time of seedling emergences under both controlled environments in laboratory and field conditions. An experiment was conducted both in the laboratory with five soil textures, three sowing depths, five soil water contents and ten constant soil temperatures, under field conditions on five sowing dates (20 February, 28 March, 19 April, 10 May, and 31 May) and three sowing depths. The results demonstrated that the Beta model can predict the time of emergence. Based on the root mean square error (RMSE), the time of emergence estimated by the Beta model was in high agreement with the time of emergence measured in the laboratory. Estimation accuracy was reduced slightly by the Beta model under field conditions. The accuracy of the Beta model was influenced by the sowing date under field conditions. So, on the first and second sowing dates (with low air temperature), the estimation of time of emergence by the model was lower and on the fourth and the fifth sowing date (with warmer air temperature), was more than the duration measured. Estimation accuracy was increased by the Beta model under field conditions using soil temperature. In conclusion, the Beta model can predict the time to emergence of sugar beet seedlings in different levels of soil texture and soil water content under field conditions, and with that, the proper planting date for sugar beet seeds to overcome weeds in different soil water content can be predicted.     

Improvement of protoplast culture protocols for Beta vulgaris L. (sugar beet)

Summary The effects of NaCl, feeder cells and the embedding of protoplasts in calcium alginate have been investigated in an attempt to improve culture conditions of recalcitrant sugar beet (Beta vulgaris L.) mesophyll protoplasts. While the use of NaCl in all instances proved detrimental to protoplast development, the other two treatments had clear beneficial effects. Minimum plating densities, necessary to sustain cell division, could be reduced to <5% (<4000 protoplasts / ml) of the control levels and plating efficiencies could be significantly enhanced by approx. 10 fold. Plants could still be regenerated from soft calli derived from mesophyll protoplasts cultured under the modified conditions at a frequency of 20–30 %. In particular, the use of alginate is considered of potentially great importance for the further application of beet protoplasts for other aims e.g. asymmetric hybridization.     

Foliar uptake of Cd by pea (Pisum sativum) and sugar beet (Beta vulgaris)

Pea ( Pisum sativum L. cv. Fenomen) and sugar beet ( Beta vulgaris L. cv. Monohill) were cultivated in nutrient media without or with 10 μM CdCl₂. Leaves of the same size and stage of development, detached or still attached to the intact plants, were submerged into redistilled water containing 1 to 250 μM CdCl₂. The uptake experiments were run for 1 to 8 h at pH 3.6 and 5.1. Cuticular transpiration rate, density of leaf and density of stomata were also measured. Percentage of open stomata was studied at different pH.
Foliar uptake of Cd into the leaf is evident since Cd is transported from the exposed part of the pea leaves, through the petioles and into the stipules, and since the Cd concentration of the leaves increases with time and external Cd concentration. The foliar uptake depends on the permeability of the cuticular membrane, which is increased by a high intrinsic Cd level, which in turn enhances the foliar uptake of Cd in sugar beet. Higher cuticular permeability in pea than in sugar beet is shown by a 2.5 times higher cuticular transpiration rate and a 4 times lower density of leaf for pea, which causes a 7 times higher foliar uptake in pea than in sugar beet. Low pH decreases the net uptake of Cd, probably by an exchange reaction in the cutin and pectin of the cuticular membrane. Stomata are not directly involved in the Cd uptake, and the differences in the sum total of stomatal aperture area per unit leaf area is not related to differences in foliar uptake of Cd. Percentage of open stomata, calculated as average of both sides of the leaves, was not affected by changes in pH: but especially at high pH. proportionally more stomata were open on the adaxial than on the abaxial side.     

Evidence for a blockwise distribution of acetyl groups onto homogalacturonans from a commercial sugar beet (Beta vulgaris) pectin

Commercial acid-extracted sugar beet pectin was extensively hydrolysed using an endo-polygalacturonase (AnPGI from Aspergillus niger or AnPGII from A. niger or FmPG from Fusarium moniliforme) in combination with Aspergillus aculeatus pectin methyl-esterase (AaPME). The homogalacturonan-derived oligogalacturonates released were quantified by high-performance anion-exchange chromatography and their structure determined by mass spectrometry. The different endo-polygalacturonases exhibited variable tolerance towards acetyl groups. AnPGI was the most active and FmPG the less. A hypothetical homogalacturonan was constructed using the AnPGI-recovered oligogalacturonates as building blocks and the validity of the model was checked taking into account FmPG observed requirements and hydrolysis products. A blockwise repartition of the acetyl groups onto sugar beet pectin homogalacturonan is proposed.     

Mapping of a chloroplast RFLP marker associated with the CMS cytoplasm of sugar beet (Beta vulgaris)

The Owen cytoplasm of male-sterile sugar beet is associated with several alterations of mitochondrial DNA and one additional HindIII site of chloroplast DNA. The region of this HindIII site has been cloned and sequenced. The site maps in a small reading frame (orf32) close to the ycf7 (orf31) gene in the petG-psbE region of chloroplast DNA. Possible functional implications of the results are discussed. The chloroplast RFLP marker described could be useful for studies on chloroplast-mitochondrial interactions, CMS of sugar beet, and the origin of the Owen cytoplasm.