Search Results (12)

Crocins are glycosylated apocarotenoids with strong coloring power and anti-oxidant, anticancer, and neuro-protective properties. We previously dissected the saffron crocin biosynthesis pathway, and demonstrated that the CsCCD2 enzyme, catalyzing the carotenoid cleavage step, shows a strong preference for the xanthophyll zeaxanthin in vitro and in bacterio. In order to investigate substrate specificity in planta and to establish a plant-based bio-factory system for crocin production, we compared wild-type Nicotiana benthamiana plants, accumulating various xanthophylls together with α- and β-carotene, with genome-edited lines, in which all the xanthophylls normally accumulated in leaves were replaced by a single xanthophyll, zeaxanthin. These plants were used as chassis for the production in leaves of saffron apocarotenoids (crocins, picrocrocin) using two transient expression methods to overexpress CsCCD2: agroinfiltration and inoculation with a viral vector derived from tobacco etch virus (TEV). The results indicated the superior performance of the zeaxanthin-accumulating line and of the use of the viral vector to express CsCCD2. The results also suggested a relaxed substrate specificity of CsCCD2 in planta, cleaving additional carotenoid substrates. Full article

Among Citrus species, the sweet orange (Citrus sinensis (L.) Osbeck) is the most important in terms of production volumes and cultivated areas. Oranges are particularly appreciated for the organoleptic characteristics and the high nutraceutical value of the fruits (thanks especially to their high content of antioxidants). Recent advances in citrus genetic and genomic resources, such as the release of the reference genomes of several sweet orange cultivars, have contributed to (i) understanding the diversification of C. sinensis and its relation with other citrus species, (ii) assessing the molecular mechanisms underlying traits of interest, (iii) identifying and characterizing the candidate genes responsible for important phenotypic traits, and (iv) developing biotechnological methods to incorporate these traits into different citrus genotypes. It has been clarified that all the genetic diversity within the sweet orange species was derived from subsequent mutations starting from a single ancestor and was derived from complex cycles of hybridization and backcrossing between the mandarin (Citrus reticulata Blanco) and the pummelo (Citrus maxima (Burm.) Merr.). This paper provides an overview of the varietal panorama together with a description of the main driving forces in present and future sweet orange breeding. In fact, for the sweet orange, as well as for other citrus species, the release of novel varieties with improved characteristics is being pursued thanks to the employment of conventional and/or innovative (molecular-based) methods. The state of the art methods together with the innovations in genomics and biotechnological tools leading to the so-called new plant breeding technologies were also reviewed and discussed. Full article

The major challenges that agriculture is facing in the twenty-first century are increasing droughts, water scarcity, flooding, poorer soils, and extreme temperatures due to climate change. However, most crops are not tolerant to extreme climatic environments. The aim in the near future, in a world with hunger and an increasing population, is to breed and/or engineer crops to tolerate abiotic stress with a higher yield. Some crop varieties display a certain degree of tolerance, which has been exploited by plant breeders to develop varieties that thrive under stress conditions. Moreover, a long list of genes involved in abiotic stress tolerance have been identified and characterized by molecular techniques and overexpressed individually in plant transformation experiments. Nevertheless, stress tolerance phenotypes are polygenetic traits, which current genomic tools are dissecting to exploit their use by accelerating genetic introgression using molecular markers or site-directed mutagenesis such as CRISPR-Cas9. In this review, we describe plant mechanisms to sense and tolerate adverse climate conditions and examine and discuss classic and new molecular tools to select and improve abiotic stress tolerance in major crops. Full article

In recent years, there have been significant developments in biotechnology, specifically regarding New Plant Breeding Techniques (NPBTs). Such advancements have been driven by the need to develop improved and more sustainable crops while reducing pesticides and fertilisers. NPBTs include a heterogeneous group of methods that allow performing plant mutations more precisely than in genetically modified (GM) technologies, saving time and effort. Although some experts consider NPBTs an opportunity for organic farming expansion, the European Court of Justice in 2018 pronounced against their use in organic farming since all plants obtained by NPBTs should follow the same regulations as Genetically Modified Organisms (GMOs). This study aims to understand consumers’ attitudes and viewpoints towards new breeding techniques. Focus groups and Q methodological approach were used to uncover consensus and divergence among organic consumers in seven selected European countries (Germany, Italy, Latvia, the Netherlands, Spain, Switzerland, United Kingdom). Results of qualitative studies suggest that organic consumers are generally hostile towards NPBTs in organic farming. Using Q methodology, three distinct factors were identified: the “Risk Averse”, the “Technological Optimists”, and the “Socially Concerned”. The results highlight that consumers’ subjective knowledge and understanding of NPBTs diverge from the discourse of NPBTs lobbyists and proponents. Full article

Castanea sativa is an important tree nut species worldwide, highly appreciated for its multifunctional role, in particular for timber and nut production. Nowadays, new strategies are needed to achieve plant resilience to diseases, climate change, higher yields, and nutritional quality. Among the new plant breeding techniques (NPBTs), the CRISPR/Cas9 system represents a powerful tool to improve plant breeding in a short time and inexpensive way. In addition, the CRISPR/Cas9 construct can be delivered into the cells in the form of ribonucleoproteins (RNPs), avoiding the integration of exogenous DNA (GMO-free) through protoplast technology that represents an interesting material for gene editing thanks to the highly permeable membrane to DNA. In the present study, we developed the first protoplast isolation protocol starting from European chestnut somatic embryos. The enzyme solution optimized for cell wall digestion contained 1% cellulase Onozuka R-10 and 0.5% macerozyme R-10. After incubation for 4 h at 25 °C in dark conditions, a yield of 4,500,000 protoplasts/mL was obtained (91% viable). The transfection capacity was evaluated using the GFP marker gene, and the percentage of transfected protoplasts was 51%, 72 h after the transfection event. The direct delivery of the purified RNP was then performed targeting the phytoene desaturase gene. Results revealed the expected target modification by the CRISPR/Cas9 RNP and the efficient protoplast editing. Full article

New plant breeding techniques (NPBTs) are seen as promising and innovative tools to achieve food security and food safety. Biotechnological innovations have great potential to address sustainable food development, and they are expected in the near future to play a critical role in feeding a growing population without exerting added pressure on the environment. There is, however, a considerable debate as to how these new techniques should be regulated and whether some or all of them should fall within the scope of EU legislation on genetically modified organisms (GMOs), despite the product obtained being free from genes foreign to the species. In the EU, the adoption of these methods does not rely only on the scientific community but requires social acceptance and a political process that leads to an improved regulatory framework. In this paper, we present the results of an online survey carried out in Italy with 700 randomly selected participants on consumer attitudes towards food obtained by NPBTs. By applying the decision tree machine learning algorithm J48 to our dataset, we identified significant attributes to predict the main drivers of purchasing such products. A classification model accuracy assessment has also been developed to evaluate the overall performance of the classifier. The result of the model highlighted the role of consumers’ self-perceived knowledge and their trust in the European approval process for NPBT, as well as the need for a detailed label. Our findings may support decision makers and underpin the development of NPBT products in the market. Full article

Two Neofusicoccumparvum isolates and a UV mutant were characterized for their phytotoxin production in vitro, their pathogenicity on grapevine, and their genome sequenced. The isolate Np-Bt67 produced high level of (-)-terremutin, but almost no (R)-mellein, and it was the most aggressive on grapevine, triggering apoplexy. Similar symptoms were not induced by purified (-)-terremutin. The isolate Bourgogne S-116 (Np-B) produced 3-fold less (-)-terremutin and high amounts of (R)-mellein, but it was less aggressive on grapevine than Np-Bt67. The UV9 mutant obtained from Np-B (NpB-UV9) no longer produced (-)-terremutin but overproduced (R)-mellein by 2.5-fold, and it was as pathogenic as its parent. NpB-UV9 differed from its parent by simple mutations in two genes (transcription factor UCR-NP2_6692, regulatory protein UCR-NP2_9007), not located neither near (R)-mellein, nor (-)-terremutin biosynthetic genes, but likely involved in the control of (-)-terremutin biosynthesis. Grapevine immunity was disturbed upon challenge with these pathogens or purified phytotoxins, leading to an upregulation of SA-dependent defenses, while (-)-terremutin interfered with host JA/ET-dependent defenses. Our results suggest that neither (-)-terremutin nor (R)-mellein alone is essential for the pathogenicity of N. parvum on grapevine, since isolate/mutant non-producing these toxins in vitro is pathogenic. However, these phytotoxins could play a quantitative role in the infection process. Full article

New Plant Breeding Techniques (NPBTs) protocols have been developed to produce new grape varieties with improved quantitative and qualitative characteristics. Reliable transformation protocols for grapes are based on the generation/induction of embryogenic callus cells that are then transformed. Varieties such as Italia have proven to be very recalcitrant to regeneration via somatic embryogenesis. In this work, the development of a protocol for improved production of embryogenic calluses is described. Two sterilization protocols were tested: (a) a lower active chlorine concentration for a longer time (LS); and (b) a higher chlorine concentration for a shorter time (HS), in combination with the absence or presence of citric acid in the growing substrate in the first growth media. The embryogenic calluses formation in Chardonnay, a cv. with a high embryogenic response, was significantly higher in presence of citric acid in the initial growing substrate regardless of the sterilization protocol. In Aglianico, a cv. with a lower embryogenic response, no significant differences were observed. Instead, in a recalcitrant cv. as Italia, we obtained a 13-fold increase in embryogenic calluses formation performing sterilization of flowers with the HS protocol compared to LS. Full article

Crop plants often have challenges of biotic and abiotic stresses, and they adapt sophisticated ways to acclimate and cope with these through the expression of specific genes. Changes in chromatin, histone, and DNA mostly serve the purpose of combating challenges and ensuring the survival of plants in stressful environments. Epigenetic changes, due to environmental stress, enable plants to remember a past stress event in order to deal with such challenges in the future. This heritable memory, called “plant stress memory”, enables plants to respond against stresses in a better and efficient way, not only for the current plant in prevailing situations but also for future generations. Development of stress resistance in plants for increasing the yield potential and stability has always been a traditional objective of breeders for crop improvement through integrated breeding approaches. The application of epigenetics for improvements in complex traits in tetraploid and some other field crops has been unclear. An improved understanding of epigenetics and stress memory applications will contribute to the development of strategies to incorporate them into breeding for complex agronomic traits. The insight in the application of novel plant breeding techniques (NPBTs) has opened a new plethora of options among plant scientists to develop germplasms for stress tolerance. This review summarizes and discusses plant stress memory at the intergenerational and transgenerational levels, mechanisms involved in stress memory, exploitation of induced and natural epigenetic changes, and genome editing technologies with their future possible applications, in the breeding of crops for abiotic stress tolerance to increase the yield for zero hunger goals achievement on a sustainable basis in the changing climatic era. Full article

It is abundantly clear that the development of gene editing technologies, represents a potentially powerful force for good with regard to human and animal health and addressing the challenges we continue to face in a growing global population. This now includes the development of approaches to modify microalgal strains for potential improvements in productivity, robustness, harvestability, processability, nutritional composition, and application. The rapid emergence and ongoing developments in this area demand a timely review and revision of the current definitions and regulations around genetically modified organisms (GMOs), particularly within Europe. Current practices within the EU provide exemptions from the GMO directives for organisms, including crop plants and micro-organisms that are produced through chemical or UV/radiation mutagenesis. However, organisms generated through gene editing, including microalgae, where only genetic changes in native genes are made, remain currently under the GMO umbrella; they are, as such, excluded from practical and commercial opportunities in the EU. In this review, we will review the advances that are being made in the area of gene editing in microalgae and the impact of regulation on commercial advances in this area with consideration to the current regulatory framework as it relates to GMOs including GM microalgae in Europe. Full article

Several papers in a Special Issue of Sustainability have recently discussed various aspects to evaluate whether organic farming and gene manipulation are compatible. A special emphasis was given to new plant breeding techniques (NPBTs). These new approaches allow the most predictable genetic alterations of crop plants in ways that the genetically modified plant is identical to a plant generated by conventional breeding. The articles of the Special Issue present the arguments pro and contra the inclusion of the plants generated by NPBTs in organic farming. Organic movements have not yet made a final decision whether some of these techniques should be accepted or banned. In my view these novel genetically manipulated (GM) crops could be used in such a way as to respect the requirements for genetically manipulated organisms (GMOs) formulated by the International Federation of Organic Movements (IFOAM). Reviewing the potential benefits of disease-resistant potatoes and bananas, it seems possible that these crops support organic farming. To this end, I propose specific requirements that the organic movements should proactively formulate as their standards to accept specific GM crops. Full article

Biotechnology is diversifying rapidly through the development and application of new approaches to genome editing and ongoing research into synthetic biology. Proponents of biotechnology are enthusiastic about these new developments and have recently begun calling for environmental movements to abandon their campaigns against Genetically Modified Organisms (GMOs) and for organic agriculture to reconsider its exclusion of Genetic Modification (GM). In this article, we begin by describing the diversity of practices that cluster under both the terms GM and organic and show that although there is a clash of different cultures of agriculture at stake, there is also a spectrum of practices existing between these two poles. Having established the terms of the debate, we then go on to analyse whether the organic movement should reconsider its position on GM in light of new plant breeding techniques (NPBTs), using the criteria highlighted as important by the International Federation of Organic Agriculture Movements (IFOAM) in their 2016 draft revised position on GMOs. Through this analysis, we suggest that given the in-context-trajectory of biotechnology development, the continued narrow framing of agricultural problems and the ongoing exclusion of important socio-economic, political and cultural dimensions, the organic movement is justified in maintaining its opposition to GM in the face of NPBTs. Full article