Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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Potatoes, pathogens and pests : effects of genetic modifi cation for plant resistance on non-target arthropods
Lazebnik, Jenny - \ 2017
University. Promotor(en): Joop van Loon; Marcel Dicke. - Wageningen : Wageningen University - ISBN 9789463431620 - 151
solanum tuberosum - potatoes - oomycetes - phytophthora infestans - genetic engineering - transgenic plants - disease resistance - risk assessment - nontarget organisms - arthropods - insect pests - herbivores - trophic levels - ecological risk assessment - greenhouse experiments - field experimentation - aardappelen - oömyceten - genetische modificatie - transgene planten - ziekteresistentie - risicoschatting - niet-doelorganismen - geleedpotigen - insectenplagen - herbivoren - trofische graden - ecologische risicoschatting - kasproeven - experimenteel veldonderzoek

Currently, fungicides are necessary to protect potato crops against late blight, Phytophthora infestans, one of the world’s most damaging crop pathogens. The introgression of plant resistance genes from wild potato species targeted specifically to the late blight pathogen into susceptible potato varieties may alleviate the environmental impact of chemical control. Genetically modified plants are subject to an environmental risk assessment, and this includes testing for risks to the non-target arthropod community associated with the crop. The thesis begins with a review about the main plant defense responses and their role in influencing sequential interactions between herbivores and plant pathogens. The experimental chapters each focus on different aspects of the interaction between potato plants (both resistant and susceptible), the target pathogen (P. infestans) and several non-target insects. With each chapter, the scope widens: from the molecular gene expression in potato leaves in response to sequential attacks, to field scale biodiversity analyses. At the molecular level, one of the main findings was that the genomic position of the Rpi-vnt1 insertion conferring resistance to P. infestans influenced potato gene expression measured in leaves, when interacting with the non-target insect pests Myzus persicae (Green peach aphid) and Leptinotarsa decemlineata (Colorado potato beetle). Insect performance differed between the resistant GM and susceptible non-GM comparator. In the following chapter, the differences in insect performance were tested across a range of conventionally bred cultivars varying in resistance to P. infestans. Differences in M. persicae performance between several cultivars greatly outweighed the differences previously detected between the GM and non-GM comparator. These results are crucial in shaping the way risk is assessed in the context of GM crops, and these results are supported in our experiments assessing effects on biodiversity with pitfall traps in the field. The third trophic level was also addressed by comparing the performance of the parasitoid Aphidius colemani reared on GM and non-GM fed aphids, both with an without exposure to P. infestans. Differences in parasitoid performance were only found on the susceptible cultivar when inoculated with P. infestans. In the last experimental chapter the risk assessment is taken to the field comparing pitfall trap catches over two years and in two countries. Different methods for statistical analysis of biodiversity data were compared to arrive at recommendations for such analysis in the framework of environmental risk assessments. Drawing on these lessons, the discussion ends with ideas for the development of protocols for environmental risk assessments in the light of expected scientific progress in agricultural biotechnology.

Bacterial cell factoriest : applying thermophiles to fuel the biobased economy
Kranenburg, Richard van - \ 2017
Wageningen : Wageningen University & Research - ISBN 9789463431750 - 19
industriële microbiologie - chemie op basis van biologische grondstoffen - biobased economy - bacteriën - genetische modificatie - thermofiele bacteriën - biomassaconversie - industrial microbiology - biobased chemistry - bacteria - genetic engineering - thermophilic bacteria - biomass conversion
The research of Bacterial Cell Factories aims to apply bacteria for production of biobased chemicals from renewable resources. The focus lies on thermophilic Gram-positives. This group of relatively unexplored thermophiles has many relevant characteristics that make them attractive as production organism for green chemicals. Development of genetic tools is a requirement for high-throughput engineering. The scientific challenge lies in exploring and exploiting the microbial physiology of the selected production organisms, involving an integrated approach of various disciplines. Successful development of such Bacterial Cell Factories is crucial for establishing the biobased economy.
Bacteriophage: from exploration to exploitation
Nobrega, Franklin L. - \ 2017
University. Promotor(en): John van der Oost, co-promotor(en): J. Azeredo; Stan Brouns. - Wageningen : Wageningen University - ISBN 9789463430524 - 338
bacteriophages - hosts - interactions - genetic engineering - methodology - screening - bacteriofagen - gastheren (dieren, mensen, planten) - interacties - genetische modificatie - methodologie - screenen

Over the past decades, bacteriophage research has revealed the abundance of phages in nature, their morphological and genomic diversity, their influence in the regulation of microbial balance in the ecosystem and their impact on the evolution of microbial diversity. Since the 1950s, phages have also played a central role in some of the most significant fundamental discoveries in biological sciences that have been crucial for the development of molecular biology. More recently, phage research has resulted in the development of genome editing tools, and it has generated the renewed interest of using phages and phage-related products as therapeutic agents. Although major progress has been made, basic understanding on phage biology is still lacking. The number of phage genes with unknown function still largely outnumber those with established roles. Therefore, further progress depends on a deeper understanding on phage biology.

The present thesis aims at developing tools to support phage research, explores the use of phages for therapeutic purposes, and expands our insights into the biology of phages. A literature review on the molecular, structural and evolutionary determinants of phage-host interaction (Chapters 1 and 2) underlines the relatively poor understanding of the subject. A great variety of structures and mechanisms of infection are being revealed, but no correlations have yet been established between these and host interaction. Furthermore, so far no evolutionary model accurately describes the coevolution of phages and bacteria. A particular interest of evolutionary studies concerns the understanding of the prevalence of broad-host range phages in natural environments, since these are rarely isolated using standard laboratory isolation procedures. Indeed, we have tried to isolate broad host range phages targeting the Escherichia coli reference collection (Chapter 3), but found narrow-host range phages to be more prevalent. Only one phage of relatively broad host range was found (S2-36s), being able to infect 14 of the 72 strains. Proteins of interest for further exploration were found, such as depolymerases and colanic acid-degrading proteins, both with potential anti-biofilm activity.

The isolation procedures against the ECOR collection proved to be challenging due to the amount of strains and samples to be evaluated. Consequently, a high-throughput methodology was developed to simplify these isolation procedures (Chapter 4). By automated monitoring of cell growth in 96-well plates it is possible to use differences in optical densities (plotted as heatmaps) between cells subjected to the samples and in control conditions to screen for the presence of phages. The method revealed an accuracy of 98% and reduced the workload by 90%. The method developed can also be used to screen for broad-host range phages or to screen collections of phages for variants or cocktails that are suitable for treating bacterial infections. A discussion is provided of the advantages and limitations of phages for therapeutic applications (Chapter 5). It is suggested that phages in their natural state cannot be used in therapeutic applications. The future of phage therapy may possibly be genome engineering for tailoring of phage properties. Subsequently, the genetic modification of phage T7 was shown to improve (2-log) the capacity of the phage to resist to the strongly acidic conditions and enzymatic challenges of the gastrointestinal tract (Chapter 6). This was achieved by modifying the phage to express a signal peptide on its capsid to which phospholipids attach forming a protective coating. The removal of the phospholipid coating using phospholipase caused reversion to the pH-sensitive phenotype of the wild-type phage. In case of orally-delivered phages, this may improve the efficacy of phage therapy.

Engineering of phage genomes can also support evolutionary studies and basic phage research, e.g. analyzing if a certain gene is essential. A strategy developed for the random recombination of phage genomes (Chapter 7) demonstrated that it is possible to create novel productive phages by combining elements of different phage families. The findings reveal an unexpected level of flexibility and adaptability of phage genomes to accommodate and re-arrange genetic information, reflecting the pre-existing evolutionary compatibility of genes from different phages. The method is further expected to serve as a platform for improving our understanding of phage gene function and importance, where the random recombination of a single phage genome may be the preferred approach.

A different approach for the therapeutic application of phages was explored. Using phage display it was possible to identify peptides targeting claudin-low breast cancer cells (Chapter 8) and osteoarthritis cells (Chapter 9) with high levels of specificity. The peptides identified may contribute to an early detection of claudin-low breast carcinomas, and to develop more individualized therapies for both breast cancer and osteoarthritis.

In summary, the work developed in this thesis has resulted in new methodologies and biological data, thereby contributing to an increased understanding of phage biology and of the opportunities for the use of phages for diagnosis and therapy.

Starch meets biotechnology : in planta modification of starch composition and functionalities
Xu, Xuan - \ 2016
University. Promotor(en): Richard Visser, co-promotor(en): Luisa Trindade. - Wageningen : Wageningen University - ISBN 9789462579200 - 169
starch - potato starch - potatoes - solanum tuberosum - plant biotechnology - biotechnology - genetic engineering - transgenic plants - modified starches - phosphate - arabidopsis thaliana - plant breeding - zetmeel - aardappelzetmeel - aardappelen - plantenbiotechnologie - biotechnologie - genetische modificatie - transgene planten - gemodificeerd zetmeel - fosfaat - plantenveredeling

Storage starch is an energy reservoir for plants and the major source of calories in the human diet. Starch is used in a broad range of industrial applications, as a cheap, abundant, renewable and biodegradable biopolymer. However, starch needs to be modified before it can fulfill the required properties for specific industrial applications. Genetic modification of starch, as a green technology with environmental and economic advantages, has attracted increasingly attention. Many achievements obtained from earlier studies have demonstrated the feasibility and potential of using this approach to produce starches with novel properties (Chapter 2).

The main objective of this research was to produce novel starches with enhanced functionalities through genetic modification, while gaining a better understanding of storage starch biosynthesis. A focus on potato was warranted as it represents a superior model system for storage starch biosynthesis studies and for the production of starches with novel properties. To this end, a number of enzymes from various sources have been expressed in potato tubers to modify starch phosphate content and polysaccharide structure, since these two characteristics have long been recognized as key features in starch properties.

To modify starch phosphate content and explore starch (de)phosphorylation, a human phosphatase enzyme named laforin, and modifications of it, were introduced into potato (Chapter 3). Interestingly, modified starches exhibited a significantly higher phosphate content rather than the expected lower phosphate content. Transcriptome analysis showed that the increase in phosphate content was a result of upregulation of starch phosphorylating genes, which revealed a compensatory response to the loss of phosphate content in potato starch. Furthermore, the increase of phosphate content in potato starch was reached to a threshold level. This was in line with the observations in the modified starches from overexpressed- Glucan water dikinase (GWD1) transgenic plants (Chapter 4). Furthermore, overexpression of two starch dikinases from Arabidopsis thaliana, glucan water dikinase 2 and 3 (AtGWD2 and AtGWD3), did not result in a significant increase in phosphate content of potato starch (Chapter 5). Taken together, these results indicated that phosphate content of potato starch is under strict control.

Morphological analysis of starch granules containing different levels of phosphate content confirmed the indispensible role of phosphate content in the normal formation of starch granules, since cracked granules were observed in the starches containing low phosphate content, while irregular bumpy shaped granules were observed in the tubers from plants containing high phosphate content. Interestingly, further analyses on the expression level of genes involved in starch metabolism and sugar-starch conversion suggested that starch phosphorylation might affect starch synthesis by controlling the carbon flux into starch while simultaneously modulating starch-synthesizing genes. Further studies are needed to confirm this finding (Chapter 4).

To produce starches with novel structures, an (engineered) 4, 6-α-glucanotransferase (GTFB) from Lactobacillus reuteri 121 was introduced into potato tubers (Chapter 6). The resulting starches showed severe changes in granule morphology, but not in starch fine structure. Transcriptome analysis revealed the existence of a self-repair mechanism to restore the regular packing of double helices in starch granules, which possibly resulted in the removal of novel glucose chains potentially introduced by the (engineered) GTFB.

This research successfully generated starches with various functionalities, including altered gelatinization characteristics (Chapter 3 and 4), improved freeze-thaw stability (Chapter 4) and higher digestibility (Chapter 6). The exploitation of relationships between starch characteristics and starch properties revealed that starch properties represent the outcome of the combined effect of many factors and are highly dependent on the genetic background in which the modification has been performed.

In conclusion, the research described in this thesis demonstrates the great potential of genetic modification in producing starches with novel properties. Meanwhile, these results revealed the presence of complex and exquisite molecular regulation mechanisms for starch biosynthesis in potato. In future research, these regulations need to be taken into account for the relational design of starch in planta. Certainly, a better understanding of the process of starch metabolism in storage organs would be a great step forward towards tailoring starch in an economically important crop such as potato.

Metabolic engineering of biosynthesis and sequestration of artemisinin
Wang, B. - \ 2016
University. Promotor(en): Harro Bouwmeester, co-promotor(en): Sander van der Krol. - Wageningen : Wageningen University - ISBN 9789462576728 - 210 p.
artemisinin - nicotiana benthamiana - arabidopsis - biosynthesis - malaria - drugs - genetic engineering - metabolism - artemisinine - biosynthese - geneesmiddelen - genetische modificatie - metabolisme

The sesquiterpenoid artemisinin (AN) is the most important medicine for the treatment of malaria in humans. The industrial production of AN still mainly depends on extraction from the plant Artemisia annua. However, the concentration of AN in A. annua is low. Although different engineering strategies have been used in both A. annua and heterologous plant and yeast production platforms, the worldwide capacity and production costs for AN are not in balance with its demand (Chapter 1). Although the genes encoding for the entire AN biosynthesis pathway (AN-PW) of the AN precursor dihydroartemisinic acid (DHAA) have been identified, the application of these genes in pathway engineering seem to be limited by lack of control over product transport and sequestration. At the onset of this thesis project there was no information on transport in the AN-PW. However, it was known that DHAA is converted into AN outside the glandular trichome cells of A. annua. Therefore, in this thesis I tried to gain more knowledge on transport within the AN-PW and the use of different metabolic engineering strategies to improve the production of AN.

At the onset of my PhD project, the AN-PW genes from two different A. annua chemotypes were compared to understand the basis of different relative activities in the two branches of the AN-PW (Chapter 2). For these assays we used transient expression in N. benthamiana. In the AN-PW, artemisinic aldehyde (AAA) is at a branch point as it can be converted to artemisinic acid (AA) by amorphadiene oxidase (AMO), or to dehydroartemisinic aldehyde (DHAAA) by artemisinic aldehyde Δ11 (13) reductase (DBR2). AA is the precursor for arteannuin B (AB) while DHAAA may be converted by a CYP71AV1 or an ALDH1 to dehydroartemisinic acid (DHAA), the precursor for AN. In this chapter we demonstrate that the CYP71AV1 from a high AN production (HAP) chemotype has reduced activity in the AB branch of the pathway compared to the CYP71AV1 from a low AN production (LAP) chemotype. In addition, we show that the relative expression levels of DBR2 and ALDH1 also affect the AN/AB chemotype. The low catalytic efficiency of AMO from the HAP chemotype may be caused by a deletion of seven amino acids at the N-terminus of the protein compared to CYP71AV1 from LAP. Ectopic expression of the AN-PW genes in N. benthamiana showed that the bulk of the PW products are modified by glycosylation and glutathione conjugations. These side reactions therefore compete with the biosynthesis flux towards the AN precursor DHAA. At this point in my thesis the ectopic expression of AN-PW genes in N. benthamiana had not yielded any AN. At a later stage it became clear that this was due to harvest of leaves at 5-7 days post agro-infiltration (dpi), while AN in N. benthamiana leaves expressing AN-PW genes only becomes detectable after 7 dpi.

Glycosylation of the bulk of the AN-PW products in N. benthamiana stresses the need for an efficient transport of (DH)AA to the outside of cells in order to escape from the glycosylation reactions. In Chapter 3, transport and sequestration of AN precursors was investigated by studying the effect of membrane transporters (PDRs) and lipid transfer proteins (LTPs). Hereto, two membrane transporters with activity towards AN-PW products were made available by the group of Prof. Marc Boutry and we isolated three LTP genes from Artemisia annua which showed expression in the glandular trichomes. In this chapter we show that AaLTP3 displays specific activity, together with AaPDR2 towards transport of (DH)AA to the apoplast in N. benthamiana. Moreover, infiltration experiments with (DH)AA in N. benthamiana leaves revealed that these compounds are rapidly taken up by the cells and that inside the cells there is a strong reverse flux in the AN-PW by conversion of (DH)AA towards (DH)AAA and (DH)AAOH. Subsequently we demonstrated that AaLTP3 has a stronger activity in keeping products in the apoplast than the AaPDR2 membrane transporter. Therefore, I suggest that by removal of (DH)AA from the cytosol through transport over the plasma membrane by AaPDR2 and subsequent sequestration in the apoplast by AaLTP3, AaLTP3 creates sink activity which prevents reflux of (DH)AA from the apoplast back into the cells. AaLTP3 therefore contributes to a directional flux through the AN-PW towards the end product (DH)AA. Finally, in this work we could also for the first time detect AN and AB in N. benthamiana leaves by extraction of necrotic leaves at 13 dpi.

Because in A. annua glandular trichome cells both the AN sesquiterpene biosynthesis pathway and the flavonoid biosynthesis pathway are active, we explored whether there is a functional interaction between these two major secondary metabolite biosynthesis pathways. In Chapter 4 we describe how we manipulate the flavonoid biosynthesis pathway in N. benthamiana leaves using the Antirhinum majus transcription factor Rosea1 (ROS) and test coexpression of ROS with AN-PW genes. The co-expression of ROS stimulates AN-PW product accumulation. Subsequent analysis indicates that this is most likely from transcriptional activation of the enzyme Mevalonate Kinase (MVK) in the mevalonate pathway, which provides precursors for the sesquiterpene biosynthesis pathway. In addition, we demonstrate that production of flavonoids competes with AN-PW product accumulation, as co-expression of AN-PW genes with ROS, but simultaneous inhibition of chalcone synthase (CHS) by a CHSRNAi construct, results in higher AN-PW product levels. However, accumulation of the end products AN and AB was not affected significantly. Finally, the combined expression of AN-PW+ROS+AaPDR2+AaLTP3+ CHSRNAi results in highest sequestration of (DH)AA in the apoplast and highest accumulation of the end products AN and AB in N. benthamiana.

During my thesis work, in a related project it was found that expression of another sesquiterpene biosynthesis gene (caryophyllene synthase; CST) in transgenic Arabidopsis resulted in higher caryophyllene emission for a transformant expressing a genomic DNA of CST, compared with a similar transformant expressing a CST cDNA described in literature. This suggested that ectopic expression of intron containing genes is more efficient than ectopic expression of cDNAs. To test whether in the context of metabolic engineering the use of genomic (intron-containing) genes is more efficient than the use of the corresponding cDNA we generated a set of stable transformed Arabidopsis lines with either genomic CST (gCST), cDNA CST (cCST), genomic amorphadiene synthesis (gADS) and cDNA ADS (cADS). In chapter 5 we show that indeed the lines with overexpression of the genomic clones yield higher levels of the anticipated products (caryophyllene or amorphadiene) than the lines with overexpression of the corresponding cDNAs. Transcript analysis showed that for gCST the increase in caryophyllene production was higher than can be explained solely by the increase in CST transcription. In the context of transient expression in N. benthamiana leaves the intron-mediated-enhancement effect was less pronounced.

In the final discussion chapter 6 I review limitations and potential solutions to metabolic engineering of the AN-PW in plants, and I discuss the impact of our findings on AN production capacity using transient expression versus natural production in A. annua. Moreover, I discuss how the finding of this thesis go beyond just insights into the AN-PW as especially the identification of the role of LTPs in sesquestration of (sesqui)terpenes into the apoplast may have an impact on the metabolic engineering efforts of many other (sesqui)terpene pathways. Because some plant hormones are also terpenoid products the newly identified role of LTPs may also have impact on a deeper understanding of hormone signalling in plants. I have already started exploring this path by generating a set of Arabidopsis plants with overexpression of different Arabidopsis LTP genes to test whether any hormone related traits are altered (Chapter 6). Preliminary results do indeed confirm a role of LTPs in endogenous plant hormone balance, something worthwhile to be further explored in future research.

De permanente groene revolutie van Swaminathan
Fresco, L.O. ; Rabbinge, R. - \ 2015
Vork 2 (2015)3. - ISSN 2352-2925 - p. 66 - 71.
green revolution - genetic engineering - human feeding - hunger - food supply - scientific research - scientists - society - fertilizers - pesticides - environmental impact - india - netherlands - potatoes - wheat - groene revolutie - genetische modificatie - humane voeding - honger - voedselvoorziening - wetenschappelijk onderzoek - wetenschappers - samenleving - kunstmeststoffen - pesticiden - milieueffect - nederland - aardappelen - tarwe
Het weekblad Time kwalificeerde hem als een van de twintig meest invloedrijke Aziaten van de twintigste eeuw: ‘The father of the Green Revolution used his skills in genetic engineering and his powers of persuasion to make famine an unfamiliar word in Asia’. Tegelijkertijd wees hij al vroeg op de gevaren van een te grote afhankelijkheid van kunstmest en bestrijdingsmiddelen en milieugevolgen daarvan. Dr. Monkombu Sambasivan Swaminathan is een fervent pleitbezorger van de Evergreen Revolution, de permanente groene revolutie. Op 7 augustus werd hij 90 jaar. Louise Fresco en Rudy Rabbinge schetsen zijn enorme betekenis voor wetenschap en samenleving.
Economic impact of the Commission's 'opt-out' proposal on the use of approved GM crops : quick assessment of the medium-term economic consequences
Hoste, R. ; Wagenberg, C.P.A. van; Wijnands, J.H.M. - \ 2015
LEI Wageningen UR (Report / LEI Wageningen UR 2015-097) - ISBN 9789086157259 - 51 p.
transgenic plants - crops - genetic engineering - soyabeans - economic impact - agricultural sector - food industry - feed industry - european union - france - germany - poland - hungary - transgene planten - gewassen - genetische modificatie - sojabonen - economische impact - landbouwsector - voedselindustrie - veevoederindustrie - europese unie - frankrijk - duitsland - polen - hongarije
The European Commission proposed the opportunity for individual EU Member States to restrict or prohibit the use of GMOs in food or feed on their territory (a national ‘opt-out’). The economic impact on individual sectors of the feed and food chain (the vegetable oil and meal industry, trade, animal feed industry, livestock sector) of a possible opt-out policy for soy by individual Member States has been assessed by LEI Wageningen UR.
A single scenario was defined in which the four countries France, Germany, Poland and Hungary choose an ‘opt-out’ policy for soy. Consequences of this switch to non-GM soy and substitutes were assessed both quantitatively and qualitatively for feed prices, for production costs for animal production, for crushing industry and for trade, with a focus on the medium term
Effecten van een verbod op het gebruik van genetisch gemodificeerde soja als veevoedergrondstof. Quick scan van de gevolgen voor Nederland
Wagenberg, C.P.A. van; Hoste, R. - \ 2015
Wageningen : LEI Wageningen UR (LEI Report 2015-109) - ISBN 9789086157143 - 26
transgenic plants - crops - genetic engineering - soyabeans - fodder - economic impact - netherlands - transgene planten - gewassen - genetische modificatie - sojabonen - veevoeder - economische impact - nederland
If the Netherlands, alongside Germany, France, Poland, and Hungary, decides to ban genetically modified (GM) soy in animal feed, the use of soy products in animal feed in these five countries will have to decrease by 40 to 50% to ensure that the EU demand for non-GM soy does not exceed the supply on the world market. The extra costs to Dutch livestock farmers over a period of 3 to 5 years as a result of the more expensive non-GM soy and alternative protein sources are estimated at between €60 and €100 million a year, with approximately 80% being borne by poultry farmers. Livestock numbers and productivity will then be maintained. A partial shift in trade flows from animal feed ingredients can be expected from import in the west of the EU - for example, through the port of Rotterdam - to intra-EU flows from production areas within the EU to consumers and via the waterway axis from regions east of the EU, such as Ukraine. Less soy will enter the EU via the Netherlands. This deficit can be offset by the increased demand for alternative protein sources, which will be partly imported from overseas. The effects on Dutch ports, the transport sector, and employment will depend on the nature of the trade flow shifts.
DNA : the recipe book for all the processes in the plant : all cells have the same generic information
Heuvelink, E. ; Kierkels, T. - \ 2015
In Greenhouses : the international magazine for greenhouse growers 4 (2015)4. - ISSN 2215-0633 - p. 12 - 13.
dna - plantenveredeling - genetische modificatie - transfer rna - messenger rna - ribosomen - eiwitten - aminozuren - enzymen - mutaties - plant breeding - genetic engineering - ribosomes - proteins - amino acids - enzymes - mutations
It’s sometimes called a blueprint: DNA, the carrier of genetic information. But the term recipe book covers it better. It explains how the plant can respond to changing conditions. Plant breeders take advantage of natural variations in DNA. Genetic modification can make their job easier.
Synthetische biologie komende jaren in het vizier
Sikkema, A. ; Martins dos Santos, V.A.P. - \ 2015
Resource: weekblad voor Wageningen UR 9 (2015)20. - ISSN 1874-3625 - p. 12 - 13.
genetische modificatie - moleculaire biologie - bacteriën - industriële microbiologie - systeembiologie - biobased economy - genetic engineering - molecular biology - bacteria - industrial microbiology - systems biology
Wageningen UR gaat de komende jaren investeren in synthetische biologie. Het is benoemd als een van de investeringsthema’s in het strategisch plan. Een jong vakgebied, waarbij onderzoekers bacteriën verbouwen en ontwerpen voor industriële toepassingen. Hoogleraar Martins dos Santos, die onlangs twee grote EU-projecten binnenhaalde, gaat een actieplan schrijven.
Voorstel voor een co-existentie monitoringsprogramma t.b.v. het naast elkaar bestaan van genetisch gemodificeerde (GG) en niet-GG teelten in toekomstige praktijksituaties. 3. Suikerbiet
Wiel, C.C.M. van de; Kok, E.J. ; Scholtens, I.M.J. ; Smulders, M.J.M. ; Lotz, L.A.P. - \ 2015
Wageningen UR - 36
akkerbouw - veldgewassen - suikerbieten - vermenging - kruisbestuiving - genetische modificatie - uitkruisen - transgene planten - nederland - arable farming - field crops - sugarbeet - mixing - cross pollination - genetic engineering - outcrossing - transgenic plants - netherlands
Beschrijving van een voorstel voor een concreet co-existentiemonitoringprogramma (CMP) voor suikerbiet dat is aangepast aan de specifieke gewaseigenschappen van suikerbiet. De gemaakte keuzen t.b.v. een pragmatische invulling van het voorgestelde CMP worden in de opvolgende hoofdstukken toegelicht op basis van de huidige stand van zaken in het wetenschappelijk onderzoek aan (trans)genverspreiding in suikerbiet. Er is nog geen ervaring met een CMP in suikerbiet, noch is er een (Europese) standaard voor. Voor het bereiken van een zo pragmatisch mogelijke aanpak wordt zoveel mogelijk aangesloten bij al bestaande evaluatiepraktijken in de suikerbietenteelt
Voorstel voor een co-existentie monitoringsprogramma t.b.v. het naast elkaar bestaan van genetisch gemodificeerde (GG) en niet-GG teelten in toekomstige praktijksituaties. 2. Aardappel
Wiel, C.C.M. van de; Kok, E.J. ; Scholtens, I.M.J. ; Smulders, M.J.M. ; Lotz, L.A.P. - \ 2015
Wageningen UR - 36
akkerbouw - aardappelen - veldgewassen - vermenging - kruisbestuiving - genetische modificatie - uitkruisen - transgene planten - nederland - arable farming - potatoes - field crops - mixing - cross pollination - genetic engineering - outcrossing - transgenic plants - netherlands
Het rapport is als volgt opgezet. Eerst wordt een voorstel voor een concreet coexistentiemonitoringprogramma (CMP) voor aardappel beschreven dat is aangepast aan de specifieke gewaseigenschappen van aardappel. De gemaakte keuzen t.b.v. een pragmatische invulling van het voorgestelde CMP worden in de opvolgende hoofdstukken toegelicht op basis van de huidige stand van zaken in het wetenschappelijk onderzoek aan (trans)genverspreiding in aardappel. Er is nog geen ervaring met een CMP in aardappel, noch is er een (Europese) standaard voor. Voor het bereiken van een zo pragmatisch mogelijke aanpak wordt zoveel mogelijk aangesloten bij al bestaande evaluatiepraktijken in de aardappelteelt. Dat betekent dat zoveel mogelijk aangesloten is bij de bestaande controlepraktijk zoals die uitgevoerd wordt door de NAK. Dat laat onverlet dat co-existentiemonitoring niet tot de staande praktijk van de NAK gerekend kan worden en dat nu ook nog niet vastligt dat de NAK deze monitoring inderdaad gaat doen. Praktische uitvoering hangt af van besluitvorming over de uiteindelijke invulling van de co-existentiemonitoring zodra GG aardappelteelt geïntroduceerd zou gaan worden.
Voorstel voor een co-existentie monitoringsprogramma t.b.v. het naast elkaar bestaan van genetisch gemodificeerde (GG) en niet GG-teelten in toekomstige praktijksituaties. 1. Maïs
Wiel, C.C.M. van de; Kok, E.J. ; Scholtens, I.M.J. ; Dolstra, O. ; Smulders, M.J.M. ; Lotz, L.A.P. - \ 2015
Wageningen UR - 33
akkerbouw - veldgewassen - maïs - genetische modificatie - vermengen - uitkruisen - transgene planten - nederland - arable farming - field crops - maize - genetic engineering - blending - outcrossing - transgenic plants - netherlands
In het rapport wordt een voorstel voor een concreet co-existentiemonitoringprogramma (CMP) voor maïs beschreven dat is aangepast aan de specifieke gewaseigenschappen van maïs. De gemaakte keuzen t.b.v. een pragmatische invulling van het voorgestelde CMP worden in de opvolgende hoofdstukken toegelicht op basis van de huidige stand van zaken in het wetenschappelijk onderzoek aan (trans)genverspreiding in maïs. Er is nog beperkte ervaring met een CMP in maïs, bijvoorbeeld in Portugal, Tsjechië en Slowakije waar Bt MON810 maïs op beperkte schaal verbouwd wordt. Er is ook geen (Europese) standaard voor een CMP, maar er is wel voor maïs als eerste een Best Practice Document door het European Co-existence Bureau (ECoB) van het JRC uitgebracht (Rizov & Rodríguez-Cerezo 2014).
Prof. Justus Wesseler over overheidsingrijpen bij nieuwe biotechnologische ontwikkelingen
Wesseler, J.H.H. - \ 2014
Wageningen UR
genetische modificatie - transgene planten - transgene organismen - biotechnologie - schade - rijst - india - economische aspecten - genetic engineering - transgenic plants - transgenic organisms - biotechnology - damage - rice - economic aspects
Overheidsregels rond biotechnologie en genetisch gemodificeerde gewassen leiden vaak tot substantieel hoge investeringskosten. Die hebben weer een lager niveau van productontwikkeling tot gevolg en een concentratie in de industrie, een herschikking van onderzoeksprioriteiten en een verschuiving van onderzoek en ontwikkeling naar landen met minder stringente regelgeving. Die trend leidde zelfs tot schade aan duurzame ontwikkeling uit oogpunt van milieu en volksgezondheid.
Biovoeding is een luxeproduct voor de middenklasse : Nederlandse voedselexpert en dwarsdenker Louise Fresco
Fresco, L.O. - \ 2014
De Tijd (België)
voedselproductie - voedselveiligheid - genetische modificatie - biologische voedingsmiddelen - biologische landbouw - intensieve veehouderij - food production - food safety - genetic engineering - organic foods - organic farming - intensive livestock farming
Binnenkort staan onze tafels weer vol exquise gerechten. Bij voorkeur gemaakt met biologische of ambachtelijke producten. Maar volgens de Nederlandse voedselexpert en dwarsdenker Louise Fresco is geïndustrialiseerde landbouw de toekomst. ‘We verlangen naar een verleden dat nooit heeft bestaan.
The discursive other dynamics in plant scientists' talk on Phytophthora with experts and the public
Mogendorff, K.G. - \ 2014
University. Promotor(en): Cees van Woerkum; Bart Gremmen, co-promotor(en): Hedwig te Molder. - Wageningen : Wageningen University - ISBN 9789462570467 - 150
communicatie - genetische modificatie - planten - technologie - discoursanalyse - openbare mening - genomica - plantkunde - wetenschappers - deskundigen - phytophthora infestans - discussie - antropologie - psychologie - communication - genetic engineering - plants - technology - discourse analysis - public opinion - genomics - botany - scientists - experts - discussion - anthropology - psychology
This dissertation investigates the interactional effects of Dutch plant science experts' talk in different interaction settings: public meetings, expert board meetings and ethnographic interviews. The main research approach deployed is discursive psychology : a methodology that focuses not on what is said but on what is accomplished with talk. The central topic of all the talk analysed in this thesis is Phytophthora Infestans: a major plant disease in staple crops that helped bring about the Irish famine in the 19th century. Phytophthora is still a large problem. To fight Phytophthora, plant experts have been developing different technologies, some of which, such as genetic modification, are met with public controversy.
Using genetic transformation to produce enhanced levels of erucic acid and novel wax esters in Crambe abyssinica
Qi, W. - \ 2014
University. Promotor(en): Richard Visser, co-promotor(en): Robert van Loo; Frans Krens. - Wageningen : Wageningen University - ISBN 9789462570368 - 117
genetische modificatie - moleculaire veredeling - moleculaire genetica - crambe abyssinica - olieleverende planten - industriële gewassen - biobased economy - erucazuur - esters - genetic engineering - molecular breeding - molecular genetics - oil plants - industrial crops - erucic acid
Crambe abyssinica (crambe) is een oliehoudend gewas dat grote potentie heeft als industrieel uitgangsmateriaal. In het onderzoek beschreven in dit proefschrift werd een solide transformatieprotocol ontwikkeld voor crambe. Om tegemoet te komen aan de eisen voor introductie in het milieu die de EU stelt aan GM gewassen en om de kans op acceptatie door het publiek te vergroten is het 'merker-vrije' genetische vector systeem dat door Wageningen UR Plant Breeding is ontwikkeld in crambe toegepast en uitgetest. Verschillende transformatie experimenten zijn uitgevoerd om het gehalte erucazuur in de zaadolie van crambe te verhogen. Ten slotte is GM crambe gebruikt als productieplatform voor hoogwaardige, industriële wasesters van het type jojoba wasesters.
Merkervrije lelies met luisresistentie : Deel 2 : Eindrapport
Krens, F.A. - \ 2014
Wageningen : Plant Research International, Wageningen UR Plant Breeding - 19
lilium - plantenveredeling - veredelingsmethoden - genetische modificatie - cultivars - resistentie van variëteiten - insectenplagen - aphis gossypii - plant breeding - breeding methods - genetic engineering - varietal resistance - insect pests
Genetisch gemodificeerde lelielijnen, die in een eerder project geproduceerd waren (PT project 12966), zijn in dit vervolgproject verder gekarakteriseerd. Het doel van project PT12966 en dit project was om GM lelies te maken die een tweetal nieuwe genen hebben ontvangen die ze resistent tegen of in ieder geval minder gevoelig zouden maken voor aantasting door bladluizen. Met als doel de introductie van resistentie tegen luizen is natuurlijk de ultieme test een resistentietoets, waarin planten blootgesteld worden aan doelinsecten, in dit geval de katoenluis, Aphis gossypii, als belager van lelies. Twee type toetsen zijn uitgevoerd aan 62 GM lelielijnen.
R gene stacking by trans- and cisgenesis to achieve durable late blight resistance in potato
Zhu, S. - \ 2014
University. Promotor(en): Evert Jacobsen; Richard Visser, co-promotor(en): Jack Vossen. - Wageningen : Wageningen University - ISBN 9789461735706 - 164
solanum tuberosum - aardappelen - phytophthora infestans - oömycota - plantenziekteverwekkende schimmels - ziekteresistentie - genen - cisgenese - transgene planten - plantenveredeling - genetische modificatie - potatoes - oomycota - plant pathogenic fungi - disease resistance - genes - cisgenesis - transgenic plants - plant breeding - genetic engineering

Among the many diseases of potato (Solanum tuberosum L.), which is the third food crop in the world after wheat and rice, late blight caused by the oomycete pathogen Phytophthora infestans, is one of the most serious diseases. In the last century, major resistance (R) genes were introgressed mainly from the wild species Solanum demissum into the cultivated potato Solanum tuberosum. However, introgression of late blight resistance genes by interspecific crosses followed by backcrosses, proved to be associated with linkage drag problems. The desired R gene is then closely linked with one or more unfavorable genes. Moreover, the obtained resistance in the varieties could be easily overcome by fast evolving virulence among P.infestans isolates. The introduction of the A2 mating type from Mexico to Europe resulted in genetically more diverse and complex P.infestans offspring, since initially only the A1 mating type existed. Therefore, new strategies for breeding varieties with durable and broad spectrum resistance needed to be developed.

Previous research indicated that varieties containing single major R genes did not show durable resistance. Therefore, the potato breeding and research community abandoned the introgression of major R genes and started breeding for horizontal resistance by combining multiple partial resistance genes. This quantitative resistance breeding approach was also not successful because the levels of resistance were too low, breeding was too complicated and the spectrum was not as broad as anticipated. Nowadays, the introgression of major R genes regained interest and two ways of resistance breeding can be distinguished: 1. molecular marker assisted resistance breeding or 2. genetic modification (GM) of existing varieties with cloned major R genes.

In this thesis, the time-saving GM approach has been investigated to achieve durable resistance against potato late blight in existing varieties by stacking of major R genes via transgenesis and cisgenesis (Chapters 2, 3, 4). These R genes are so called cisgenes and are unmodified copies of genes from the same or crossable species, harboring their own promoter and terminator sequences.

The main difference between cisgenesis and transgenesis is the resulting (end) product. The end products for the latter case are transformants, which contain transgenes, that can come from a very different species, such as the selection marker gene nptII coding for antibiotic resistance from bacteria. However, the end products of cisgenesis, called cisformants, only harbor cisgenes (which are natural genes from the same or crossable species). These cisformants are selected by PCR for the presence of R gene(s) and for the absence of vector backbone sequences. In our study, functionality of the individual R genes, in trans- and cisformants containing stacked R genes, was determined by detached leaf assays (DLA) using avirulent isolates and by agro-infiltration with Avr genes matching every single R gene. Their foliar resistance was also tested in the field, and their resistance in tubers was tested in the lab.

In order to ensure durability, an accurate and robust system must be available to monitor virulence in P.infestans populations. Differential sets with plants containing single R genes are important and developed in many crops in order to facilitate both resistance breeding and genetic research on pathogen populations in different locations worldwide. The existing conventional differential potato set of Mastenbroek was updated and a start was made to develop a GM differential set with cloned R genes in individual transformants of cv Desiree (Chapter 5).

In Chapter 2, R genes with broad and complementary resistance spectrum were selected as a first step for R gene stacking. Selection for these R genes was performed using DLA with 44 selected late blight isolates. Out of four R genes (Rpi-sto1, Rpi-vnt1.1, Rpi-blb3, and R3a), three were selected for stacking experiments, Rpi-sto1 from S. stoloniferum, Rpi-vnt1.1 from S. venturii and Rpi-blb3 from S. bulbocastanum. Cv Desiree transformants containing these three single R genes conferred resistance to 40, 43 and 37 out of 44 isolates, respectively. The R3a containing transformant conferred resistance to only five out of 44 isolates. These three broad spectrum R genes were then combined in one binary vector pBINPLUS containing nptII as kanamycin resistance marker. Transformants containing nptII and the three R genes showed foliar resistance in DLA against two isolates PIC99189 (avrsto1, Avrvnt1, avrblb3) and EC1 (Avrsto1, avrvnt1, Avrblb3). Furthermore, the functions of these three individualR genes were confirmed using the cross reacting Avr genes from the pathogen, since no isolates were available to distinguish the function of each R gene individually due to the broad resistance spectrum. The resistance spectrum of transformants containing the three R genes Rpi-sto1, Rpi-vnt1.1 and Rpi-blb3 showed after DLA the expected sum of resistance spectrum from all three individual R genes and no indications for epistatic effects were observed (Chapter 2). These triple R genes containing transformants showed also full resistance in the field after inoculation with IPO-C (Avrsto1, Avrvnt1, avrblb3) both in 2011 and 2012. Furthermore, these three R genes were inherited to the next generation as a cluster and retained their functionality after crossing. Generally, resistance in tubers of these plants showed also the summed spectrum of all individual R genes in both generations, as was the case in the foliar resistance test. It was remarkable that transgenic Desiree plants, harboring Rpi-sto1 or Rpi-blb3,showed increased resistance in tubers, while their functional homologs Rpi-blb1 and R2, did not show resistance in tubers of conventionally bred materials. The integration of T-DNA borders and vector backbone sequences was also investigated. Around 45% of the triple R gene containing transformants harbored one or two T-DNA copies, without the integration of T-DNA borders and vector backbone (Chapter 3).

The introduction of multiple R genes was also applied to produce cisformants, plants containing only cisgenes. Three approaches were taken: 1) two cisgenes were introduced through one marker free transformation vector, 2) two cisgenes were introduced through two separate marker free vectors by co-transformation, 3) co-transformation of two vectors, one only containing nptII, and the other one is a marker free transformation vector harboring three cisgenes. This co-transformation was followed by sexual crossing to remove selection marker nptII. All three approaches were successful in the production of cisformants. The first approach produced a high percentage (73%) of cisformants but, in contrast to transgenic plants, the percentage of plants showing full resistance in DLA was relatively low (42%). The second approach produced only 4% of cisformants with stacked R genes, due to the high incidence of vector backbone sequence integration from two vectors used for co-transformation. All transformants obtained by the third approach showed full late blight resistance, which was very efficient compared to the first two approaches. This must be due to the use of the nptII selection marker. After crossing, the integration of both T-DNAs appeared to be unlinked in all tested transformants. Therefore, cisformants with active R genes could be obtained. The resistance level in tubers of cisformants was more frequently sufficient in plants with integration of two or more T-DNA copies, as it was also observed in the triple R gene transformants (Chapter 3). Not only the R genes from cisformants obtained using the third approach but also the cisformants from the first approach showed clustered inheritance in a crossing population, while the R genes segregated independently in the crossing population from a cisformant obtained using the second approach (Chapter 4).

The potato late blight differential set is used to characterize the virulence of P.infestans isolates, consisting of eleven plants which are expected to represent eleven different late blight R genes. Most differential plants were found to be susceptible to current late blight isolates, with the exception of the MaR8 and MaR9 plants. It had already been described that additional R genes were present in some members of this differential set. In Chapter 5, all eleven differential plants were tested for a hypersensitive reaction towards seven Avr genes. Only in three differential plants (MaR1, MaR2 and MaR4) no additional R genes were found, while for example MaR3,MaR8 andMaR9 contained multiple R genes. The conventional differential set was extended with F1 and BC1 segregants harboring a reduced number of these R genes and potentially containing only one R gene (R3a, R3b, R8 or R9, respectively) and with plants containing recently cloned R genes (Rpi-blb3, Rpi-sto1, Rpi-blb1, Rpi-pta1, Rpi-blb2, Rpi-vnt1.1 and Rpi-chc1). A disadvantage of the (extended) conventional differential set is that their genetic background is different which is complicating the use of this set. Moreover, for none of the extended differential plants it can be ruled out that different additional R genes are present. Therefore, a GM differential set consisting of ten transformants of cv Desiree, each harboring a single R gene was compiled. This GM differential set is more reliable for characterization of P.infestans isolates and for the functional test of individual R genes, due to the isogenic background. As a proof of concept, the conventional and the GM differential sets were compared using recently collected isolates from Dutch fields in detached leaf assays. It was found that plants containing Rpi-blb3, Rpi-blb1, Rpi-chc1, R8, R9, Rpi-vnt1.1 and Rpi-blb2 showed a broader resistance spectrum as compared to R1, R3a, R3b andR4. Furthermore, the application of the GM differential set to monitor virulence towards the different R genes in local late blight populations using trap fields was investigated. The extended conventional and the GM differential sets are on continuously growing lists, which can be in the future updated with better performing, genetically more isogenic plants harboring novel R genes, or when new R genes are transformed into cv Desiree.

In the general discussion (chapter 6), related topics from different experimental chapters are discussed simultaneously, some additional experimental data are provided and a broader view on the research area is given.

In summary, five main conclusions can be drawn from this work: 1. broad spectrum resistance in leaf and tuber with stable inheritance can be achieved by gene stacking via transgenesis and cisgenesis; 2. The frequency of cisformants with sufficient resistance at foliage and tuber level is lower than in transformants; 3. Avr genes are highly needed to test for functionality of all stacked R genes in trans- or cisformants; 4. the GM differential set can be used to accurately characterize P.infestans isolates and to assess the employability of certain R genes in particular geographic locations; and 5. genetic transformation is a unique way to improve existing susceptible potato varieties such as the cvs Bintje and Russet Burbank which are grown at relatively large areas worldwide.

Algen: de groene belofte
Lamers, P.P. ; Buiter, R. ; Hoekstra, W.P.M. - \ 2013
Den Haag : Stichting Bio-Wetenschappen en Maatschappij (Cahier / Bio-Wetenschappen en Maatschappij 3) - ISBN 9789073196711 - 72
algen - technologie - aquatische ecologie - zeewierproducten - zeeproducten - biobrandstoffen - algenteelt - genetische modificatie - biobased economy - algae - technology - aquatic ecology - seaweed products - marine products - biofuels - algae culture - genetic engineering
Algen vervullen niet alleen een essentiële rol in het water, ze staan ook in toenemende belangstelling van de technologie. Vliegtuigen zouden erop kunnen vliegen, kweekvis en andere dieren kunnen ermee worden gevoed. Algen lijken een bron van vele mooie producten en toepassingen, maar de belofte is nog geen praktijk. In dit cahier belichten wetenschappers uit het algenonderzoek wat de rol van algen in de natuur is, wat de potentie is van algen als producenten van biobrandstoffen of andere producten, en vooral wat er nog moet gebeuren voor de belofte realiteit wordt.
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