Metabolism and roles of stilbenes in plants

The plant cuticle and cell wall separate microbial pathogens from the products of plant metabolism. While microbial pathogens try to breach these barriers for colonization, plants respond to attempted penetration by a battery of wall-associated defense reactions. Successful pathogens circumvent or suppress plant nonself recognition and basal defense during penetration and during microbial reproduction. Additionally, accommodation of fungal infection structures within intact cells requires host reprogramming. Recent data highlight that both early plant defense to fungal penetration and host reprogramming for susceptibility can function at the host cell periphery. Genetic evidence has also widened our understanding of how fungal pathogens are restricted during penetration at the plant cell wall. This review summarizes the current view of how plants monitor and model their cell periphery during interaction with microbial invaders.

Cell Wall–Associated Mechanisms of Disease Resistance and Susceptibility

The broad applications of somatic embryogenesis, both in basic and applied research, have stimulated studies on the determination of in vitro conditions for the induction of somatic embryos and their conversion into plants. As a result, efficient protocols on SE induction and plant regeneration have recently become available for many plant species, including Arabidopsis thaliana (L.) Heynh., a model plant in genetics and embryogenesis. Studies on factors controlling in vitro plant morphogenesis are highly desirable not only for the development of improved regeneration systems, but also for the analysis of molecular mechanisms underlying plant embryogenesis. This review focuses on the conditions influencing the induction of embryogenic potential in in vitro cultured plant cells. The roles of explant type, endo- and exogenous plant growth regulators and stress factors in the induction of somatic embryogenesis are especially emphasized. Possible mechanisms by which different factors induce or modify embryogenic competence in cultured plant cells are also discussed. Since the production of genetically solid and true-to-type plants is desired, especially for transformation and micropropagation practice, the problem of the genetic characteristics of regenerants, in terms of their chimerism and somaclonal variation, is discussed in some detail. Special consideration is given to A. thaliana– a major model plant species for classical genetics and genomics. Recent availability of efficient embryogenic cultures in this organism makes it possible to benefit from advanced genomic research of Arabidopsis to study plant embryogenesis on the molecular level.

Factors influencing somatic embryogenesis induction and plant regeneration with particular reference to Arabidopsis thaliana (L.) Heynh

A 1.1 kb rice genomic DNA fragment, containing a chitinase gene under the control of the CaMV 35S promoter, was cloned into the rice transformation vector pGL2. After transformation of Indica rice protoplasts in the presence of polyethyleneglycol, plants were regenerated. The presence of the chimeric chitinase gene in T0 and T1 transgenic rice plants was detected by Southern blot analysis. Western blot analysis of transgenic plants and their progeny revealed the presence of two proteins with apparent molecular weights of 30 and 35 kDa that reacted with the chitinase antibody. Progeny from the chitinase-positive plants were tested for their resistance to the sheath blight pathogen, Rhizoctonia solani. The degree of resistance displayed by the transgenic plants to this pathogen correlated with the level of chitinase expression.

Genetic engineering of rice for resistance to sheath blight and other agronomic characters

Genes encoding pathogenesis-related (PR-) proteins isolated from a cDNA library of Fusarium graminearum-infected wheat spikes of scab-resistant cultivar 'Sumai-3' were transformed into susceptible spring wheat, 'Bobwhite' using a biolistic transformation protocol, with the goal of enhancing levels of resistance against scab. Twenty-four putative transgenic lines expressing either a single PR-protein gene or combinations thereof were regenerated. Transgene expression in a majority of these lines (20) was completely silenced in the T(1) or T(2) generations. Four transgenic wheat lines showed stable inheritance and expression of either a single transgene or transgene combinations up to four generations. One line co-expressing a chitinase and beta-1,3-glucanase gene combination, when bioassayed against scab showed a delay in the spread of the infection (type II resistance) under greenhouse conditions. This line and a second transgenic line expressing a rice thaumatin-like protein gene (tlp) which had moderate resistance to scab in previous greenhouse trials, along with susceptible and resistance checks were evaluated for resistance to scab under field conditions. None of the transgenic lines had resistance to scab in the field under conditions of strong pathogen, suggesting these plants lacked effective resistance to initial infection (type I resistance) under these conditions. As far as is known, this is the first report of field evaluation of transgenic wheat expressing genes for PR-proteins against disease resistance.

https://www.k-state.edu/wgrc/Publications/2003/7728.pdf

Greenhouse and field testing of transgenic wheat plants stably expressing genes for thaumatin‐like protein, chitinase and glucanase against Fusarium graminearum

Three cDNAs encoding the antifungal protein Ag-AFP from the fungus Aspergillus giganteus, a barley class II chitinase and a barley type I RIP, all regulated by the constitutive Ubiquitin1 promoter from maize, were expressed in transgenic wheat. In 17 wheat lines, stable integration and inheritance of one of the three transgenes has been demonstrated over four generations. The formation of powdery mildew (Erysiphe graminis f. sp. tritici) or leaf rust (Puccinia recondita f. sp. tritici) colonies was significantly reduced on leaves from afp or chitinase II- but not from rip I-expressing wheat lines compared with nontransgenic controls. The increased resistance of afp and chitinase II lines was dependent on the dose of fungal spores used for inoculation. Heterologous expression of the fungal afp gene and the barley chitinase II gene in wheat demonstrated that colony formation and, thereby, spreading of two important biotrophic fungal diseases is inhibited approximately 40 to 50% at an inoculum density of 80 to 100 spores per cm 2 .

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Heterologous Expression of Genes Mediating Enhanced Fungal Resistance in Transgenic Wheat

Expression of transgenic stilbene synthases in wheat causes the accumulation of unknown stilbene derivatives with antifungal activity

We report here an in vitro culture system that provides reliable, highly efficient regeneration from immature embryos of pearl millet [Pennisetum glaucum (L.) R. Br.] and sorghum [Sorghum bicolor (L.) Moench]. Immature embryos were isolated 10–20 days after pollination and cultured on various L3 media. The influence of different parameters during the callus induction phase was examined with respect to the regeneration rate: (1) the concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) and various cytokinins; (2) the addition of AgNO3; (3) the use of maltose or sucrose as a carbon source. Modifications in the phytohormones alone resulted in the regeneration of fertile sorghum plants at high efficiency. Significant increases in the regeneration rates of pearl millet genotypes were achieved by the combination of sucrose as a carbon source and silver nitrate as a potential ethylene inhibitor.

Efficient in vitro plant regeneration from immature zygotic embryos of pearl millet [Pennisetum glaucum (L.) R. Br.] and Sorghum bicolor (L.) Moench

Four different pearl millet breeding lines were transformed and led to the regeneration of fertile transgenic plants. Scutellar tissue was bombarded with two plasmids containing the bar selectable marker and the β-glucuronidase reporter gene (gus or uidA) under control of the constitutive CaMV 35S promoter or the maize Ubiquitin1 promoter (the CaMV 35S is not a maize promoter). For the delivery of the DNA-coated microprojectiles, either the particle gun PDS 1000/He or the particle inflow gun was used. The calli and regenerants were selected for their resistance to the herbicide Basta (glufosinate ammonium) mediated by the bar gene. Putative transformants were screened for enzyme activity by painting selected leaves or spraying whole plants with an aqueous solution of the herbicide Basta and by the histochemical GUS assay using cut leaf segments. PCR and Southern blot analysis of genomic DNA indicated the presence of introduced foreign genes in the genomic DNA of the transformants. Five regenerated plants represent independent transformation events and have been grown to maturity and set seed. The integration of the bar selectable and the gus reporter gene was confirmed by genomic Southern blot analysis in all five plants. All five plants had multiple integrations of both marker genes. To date, the T1 progeny of three out of four lines generated by the PDS particle gun shows co-segregating marker genes, indicating an integration of the bar and the gus gene at the same locus in the genome.

Transgenic and herbicide resistant pearl millet (Pennisetum glaucum L.) R.Br. via microprojectile bombardment of scutellar tissue

Molecular breeding of wheat in the sense of genetic engineering will be the subject of this paper It will review actual strategies for engineering fungal resistance in crops An efficient method for genetic transformation of wheat like the biolistic method using immature embryos as target tissue — together with genes that directly or indirectly inhibit the pathogen’s spread — form the basis for the development of transgenic wheat plants with improved fungal resistance

Klaus H. Oldach

Papers

Heterologous Expression of Genes Mediating Enhanced Fungal Resistance in Transgenic Wheat

Expression of transgenic stilbene synthases in wheat causes the accumulation of unknown stilbene derivatives with antifungal activity

Efficient in vitro plant regeneration from immature zygotic embryos of pearl millet [Pennisetum glaucum (L.) R. Br.] and Sorghum bicolor (L.) Moench

Transgenic and herbicide resistant pearl millet (Pennisetum glaucum L.) R.Br. via microprojectile bombardment of scutellar tissue

Molecular Breeding for Improved Biotic Stress Resistance