Efficient DNA isolation from moroccan arar tree [Tetraclinis articulata (Vahl) Masters] leaves and optimization of the rapd-pcr molecular technique

Efficient DNA isolation from Moroccan Arar tree [Tetraclinis articulata (Vahl) Masters] leaves and optimization of the RAPD-PCR molecular technique. Molecular genetic analysis of Arar tree [Tetraclinis articulata (Vahl) Masters] is often limited by the availability of fresh tissue and an efficient and reliable protocol for high quality genomic DNA extraction. In this study, two DNA extraction protocols were specifically developed for extracting high quality genomic DNA from Arar tree leaves: modified QIAgen DNA Kit and protocol developed by Ouenzar et al. (1998). DNA yield and purity were monitored by gel electrophoresis and by determining absorbance at UV (A260/A280 and A260/A230). Both ratios were between 1.7 and 2.0, indicating that the presence of contaminating metabolites was minimal. The DNA yield obtained ranged between 20 to 40 μg/g of plant materiel. The Ouenzar and collaborators protocol gave higher yield but was more time consuming compared to QIAgen Kit. However, both techniques gave DNA of good quality that is amenable to RAPD-PCR reactions. Additionally, restriction digestion and PCR analyses of the obtained DNA showed its compatibility with downstream applications. Randomly Amplified Polymorphic DNA profiling from the isolated DNA was optimized to produce scorable and clear amplicons. The presented protocols allow easy and high quality DNA isolation for genetic diversity studies within Arar tree

The 8D05 Parasitism Gene of Meloidogyne incognita Is Required for Successful Infection of Host Roots

Parasitism genes encode effector proteins that are secreted through the stylet of root-knot nematodes to dramatically modify selected plant cells into giant-cells for feeding. The Mi8D05 parasitism gene previously identified was confirmed to encode a novel protein of 382 amino acids that had only one database homolog identified on contig 2374 within the Meloidogyne apla genome. Mi8D05 expression peaked in M. incognita parasitic second-stage juveniles within host roots and its encoded protein was limited to the subventral esophageal gland cells that produce proteins secreted from the stylet. Constitutive expression of Mi8D05 in transformed Arabidopsis thaliana plants induced accelerated shoot growth and early flowering but had no visible effects on root growth. Independent lines of transgenic Arabidopsis that expressed a double-stranded RNA complementary to Mi8D05 in host-derived RNA interference (RNAi) tests had up to 90% reduction in infection by M. incognita compared with wild-type control plants, suggesting that Mi8D05 plays a critical role in parasitism by the root-knot nematode. Yeast two-hybrid experiments confirmed the specific interaction of the Mi8D05 protein with plant aquaporin tonoplast intrinsic protein 2 (TIP2) and provided evidence that the Mi8D05 effector may help regulate solute and water transport within giant-cells to promote the parasitic interaction

Plant-driven selection of microbes in the rhizosphere and plant-microbe feedbacks

  L’effet des plantes sur les microorganismes du sol et les rétroactions entre les microbes et les plantes sont devenus le focus de beaucoup de recherche. Les dernières avancées dans le domaine des interactions plantes-microbes montrent que les plantes sont capables de façonner le microbiome de leur rhizosphère via des mécanismes divers. Dans cette revue, nous rassemblons des résultats de multiples études sur le rôle des plantes dans l’altération de la structure et des fonctions des microorganismes au niveau de la rhizosphère. En outre, nous discutons les rôles de différentes substances phytochimiques dans la médiation de ces effets. Finalement, nous soulignons que l’enrichissement sélectif de microorganismes spécifiques a aussi bien des rétroactions négatives, avec l’accumulation de pathogènes dans la rhizosphère, que positives, suite au recrutement de microflore bénéfique. Une meilleure compréhension des mécanismes qui régissent la sélection de microorganismes ayant des rétroactions positives ouvrira de nouvelles possibilités pour l’amélioration de la production agricole. Mots-clés: Plantes, microorganismes, rhizosphère, rétroactions.Plant impacts on soil microbial communities and plant-microbe feedbacks have become the focus of much research. Recent advances in plant-microbe interactions investigations show that plants are able to shape their rhizosphere microbiome through diverse mechanisms. In this review, we gather findings from across multiple studies on the role of plants in altering the structure and functions of microbes in the rhizosphere. In addition, we discuss the roles of diverse phytochemicals in mediating these effects. Finally, we highlight that selective enrichment of specific microorganisms in the rhizosphere has either negative feedbacks, with pathogen accumulation in the rhizosphere; or, perhaps most importantly, positive feedbacks as a result of the recruitment of a beneficial microflora. Insights into the mechanisms that underpin plant selection of microbial communities with positive feedbacks will provide new opportunities to increase crop production. Keywords: Plants, microbes, rhizosphere, feedbacks

Plant-driven selection of microbes in the rhizosphere and plant-microbe feedbacks

  L’effet des plantes sur les microorganismes du sol et les rétroactions entre les microbes et les plantes sont devenus le focus de beaucoup de recherche. Les dernières avancées dans le domaine des interactions plantes-microbes montrent que les plantes sont capables de façonner le microbiome de leur rhizosphère via des mécanismes divers. Dans cette revue, nous rassemblons des résultats de multiples études sur le rôle des plantes dans l’altération de la structure et des fonctions des microorganismes au niveau de la rhizosphère. En outre, nous discutons les rôles de différentes substances phytochimiques dans la médiation de ces effets. Finalement, nous soulignons que l’enrichissement sélectif de microorganismes spécifiques a aussi bien des rétroactions négatives, avec l’accumulation de pathogènes dans la rhizosphère, que positives, suite au recrutement de microflore bénéfique. Une meilleure compréhension des mécanismes qui régissent la sélection de microorganismes ayant des rétroactions positives ouvrira de nouvelles possibilités pour l’amélioration de la production agricole. Mots-clés: Plantes, microorganismes, rhizosphère, rétroactions.Plant impacts on soil microbial communities and plant-microbe feedbacks have become the focus of much research. Recent advances in plant-microbe interactions investigations show that plants are able to shape their rhizosphere microbiome through diverse mechanisms. In this review, we gather findings from across multiple studies on the role of plants in altering the structure and functions of microbes in the rhizosphere. In addition, we discuss the roles of diverse phytochemicals in mediating these effects. Finally, we highlight that selective enrichment of specific microorganisms in the rhizosphere has either negative feedbacks, with pathogen accumulation in the rhizosphere; or, perhaps most importantly, positive feedbacks as a result of the recruitment of a beneficial microflora. Insights into the mechanisms that underpin plant selection of microbial communities with positive feedbacks will provide new opportunities to increase crop production. Keywords: Plants, microbes, rhizosphere, feedbacks

The interaction of the novel 30C02 cyst nematode effector protein with a plant β-1,3-endoglucanase may suppress host defence to promote parasitism

Phytoparasitic nematodes secrete an array of effector proteins to modify selected recipient plant cells into elaborate and essential feeding sites. The biological function of the novel 30C02 effector protein of the soybean cyst nematode, Heterodera glycines, was studied using Arabidopsis thaliana as host and the beet cyst nematode, Heterodera schachtii, which contains a homologue of the 30C02 gene. Expression of Hg30C02 in Arabidopsis did not affect plant growth and development but increased plant susceptibility to infection by H. schachtii. The 30C02 protein interacted with a specific (AT4G16260) host plant β-1,3-endoglucanase in both yeast and plant cells, possibly to interfere with its role as a plant pathogenesis-related protein. Interestingly, the peak expression of 30C02 in the nematode and peak expression of At4g16260 in plant roots coincided at around 3–5 d after root infection by the nematode, after which the relative expression of At4g16260 declined significantly. An Arabidopsis At4g16260 T-DNA mutant showed increased susceptibility to cyst nematode infection, and plants that overexpressed At4g16260 were reduced in nematode susceptibility, suggesting a potential role of host β-1,3-endoglucanase in the defence response against H. schachtii infection. Arabidopsis plants that expressed dsRNA and its processed small interfering RNA complementary to the Hg30C02 sequence were not phenotypically different from non-transformed plants, but they exhibited a strong RNA interference-mediated resistance to infection by H. schachtii. The collective results suggest that, as with other pathogens, active suppression of host defence is a critical component for successful parasitism by nematodes and a vulnerable target to disrupt the parasitic cycle

First draft genome assembly of the Argane tree (Argania spinosa)

Background: The Argane tree (Argania spinosa L. Skeels) is an endemic tree of southwestern Morocco that plays an important socioeconomic and ecologic role for a dense human population in an arid zone. Several studies confirmed the importance of this species as a food and feed source and as a resource for both pharmaceutical and cosmetic compounds. Unfortunately, the argane tree ecosystem is facing significant threats from environmental changes (global warming, over-population) and over-exploitation. Limited research has been conducted, however, on argane tree genetics and genomics, which hinders its conservation and genetic improvement. Methods: Here, we present a draft genome assembly of A. spinosa. A reliable reference genome of A. spinosa was created using a hybrid de novo assembly approach combining short and long sequencing reads. Results: In total, 144 Gb Illumina HiSeq reads and 7.2 Gb PacBio reads were produced and assembled. The final draft genome comprises 75 327 scaffolds totaling 671 Mb with an N50 of 49 916 kb. The draft assembly is close to the genome size estimated by k-mers distribution and covers 89% of complete and 4.3 % of partial Arabidopsis orthologous groups in BUSCO. Conclusion: The A. spinosa genome will be useful for assessing biodiversity leading to efficient conservation of this endangered endemic tree. Furthermore, the genome may enable genome-assisted cultivar breeding, and provide a better understanding of important metabolic pathways and their underlying genes for both cosmetic and pharmacological purposes

First draft genome assembly of the Argane tree (Argania spinosa) [version 2; peer review: 2 approved]

BACKGROUND : The Argane tree (Argania spinosa L. Skeels) is an endemic tree of mid-western Morocco that plays an important socioeconomic and ecologic role for a dense human population in an arid zone. Several studies confirmed the importance of this species as a food and feed source and as a resource for both pharmaceutical and cosmetic compounds. Unfortunately, the argane tree ecosystem is facing significant threats from environmental changes (global warming, over-population) and over-exploitation. Limited research has been conducted, however, on argane tree genetics and genomics, which hinders its conservation and genetic improvement. METHODS : Here, we present a draft genome assembly of A. spinosa. A reliable reference genome of A. spinosa was created using a hybrid de novo assembly approach combining short and long sequencing reads. RESULTS : In total, 144 Gb Illumina HiSeq reads and 7.6 Gb PacBio reads were produced and assembled. The final draft genome comprises 75 327 scaffolds totaling 671 Mb with an N50 of 49 916 kb. The draft assembly is close to the genome size estimated by k-mers distribution and covers 89% of complete and 4.3 % of partial Arabidopsis orthologous groups in BUSCO. CONCLUSION : The A. spinosa genome will be useful for assessing biodiversity leading to efficient conservation of this endangered endemic tree. Furthermore, the genome may enable genome-assisted cultivar breeding, and provide a better understanding of important metabolic pathways and their underlying genes for both cosmetic and pharmacological.DATA AVAILABILITY: All of the A. spinosa datasets can be retrieved under BioProject accession number PRJNA294096: http://identifiers.org/ bioproject:PRJNA294096. The raw reads are available at NCBI Sequence Reads Archive under accession number SRP077839: http://identifiers.org/insdc.sra:SRP077839. The complete genome sequence assembly project has been deposited at GenBank under accession number QLOD00000000: http://identifiers. org/ncbigi/GI:1408199612. Data can also be retrieved via the International Argane Genome Consortium (IAGC) website: http://www.arganome.org.https://f1000research.compm2021BiochemistryGeneticsMicrobiology and Plant Patholog

Efficient DNA isolation from moroccan arar tree [Tetraclinis articulata (Vahl) Masters] leaves and optimization of the rapd-pcr molecular technique. Extraction efficace de l’ADN des feuilles du thuya de berberie (Tetraclinis articulata (Vahl) masters) et optimisation de la technique moléculaire rapd-pcr

 English.  Molecular genetic analysis of Arar tree [Tetraclinis articulata (Vahl) Masters] is often limited by the availability of fresh tissue and an efficient and reliable protocol for high quality genomic DNA extraction. In this study, two DNA extraction protocols were specifically developed for extracting high quality genomic DNA from Arar tree leaves: modified QIAgen DNA Kit and protocol developed by Ouenzar et al. (1998). DNA yield and purity were monitored by gel electrophoresis and by determining absorbance at UV (A260/A280 and A260/A230). Both ratios were between 1.7 and 2.0, indicating that the presence of contaminating metabolites was minimal. The DNA yield obtained ranged between 20 to 40 µg/g of plant materiel. The Ouenzar and collaborators protocol gave higher yield but was more time consuming compared to QIAgen Kit. However, both techniques gave DNA of good quality that is amenable to RAPD-PCR reactions.Additionally, restriction digestion and PCR analyses of the obtained DNA showed its compatibility with downstream applications. Randomly Amplified Polymorphic DNA profiling from the isolated DNA was optimized to produce scorable and clear amplicons. The presented protocols allow easy and high quality DNA isolation for genetic diversity studies within Arar tree.Français.  Les analyses en génétique moléculaire chez le thuya de Berberie [Tetraclinis articulata (Vahl) Masters] sont souvent limitées par la disponibilité du matériel végétal frais et le temps nécessaire pour l’extraction l’ADN ainsi que par sa qualité. Dans cette étude, deux protocoles d’extraction, à partir des feuilles du thuya, de l’ADN génomique de haute qualité, ont été développés : le Kit Qiagen et le protocole mis au point par Ouenzar et al. (1998) modifiés. La qualité et la quantité de l’ADN sont évaluées par électrophorèse sur gel d’agarose et par la mesure de l’absorbance en UV à (A260/A280) et (A260/A230). Ces deux rapports varient entre 1,7 et 2,0 indiquant la faible fréquence des métabolites contaminants. Le rendement d’ADN varie entre 20 et 40 µg/g du matériel végétal. Le protocole de Ouenzar et collaborateurs donne le meilleur rendement d’ADN mais nécessite plus de temps. Néanmoins, les deux protocoles donnent un ADN de bonne qualité utilisable dans les réactions RAPD-PCR. En outre, la restriction enzymatique et l’analyse PCR de l’ADN obtenu ont montré sa compatibilité avec les applications moléculaires ultérieures. Les paramètres intervenant dans les réactions RAPD ont été optimisés. Les protocoles présentés permettent l’extraction facile de l’ADN de haute qualité nécessaire pour des études de la diversité génétique au sein du thuya

Modulation of Arabidopsis Flavonol Biosynthesis Genes by Cyst and Root-Knot Nematodes

Although it is well established that flavonoid synthesis is induced in diverse plant species during nematode parasitism, little is known about the regulation of genes controlling flavonol biosynthesis during the plant–nematode interaction. In this study, expression of the Arabidopsis thaliana flavonol-specific transcription factor, AtMYB12, the flavonol synthase genes, AtFLS1, 2, 3, 4, and 5, and the gene encoding the central flavonoid enzyme, chalcone synthase (AtCHS), were examined in plant roots during infection by Heterodera schachtii (sugar beet cyst) and Meloidogyne incognita (root-knot) nematodes. These experiments showed that AtMYB12 was transiently upregulated at 9 dpi in syncytia associated with sugar beet cyst nematode infection and that an Atmyb12-deficient line was less susceptible to the parasite. This suggests that, rather than contributing to plant defense, this gene is essential for productive infection. However, the AtCHS and AtFLS1 genes, which are controlled by AtMYB12, did not exhibit a similar transient increase, but rather were expressly downregulated in syncytia relative to adjacent uninfected root tissue. Genetic analyses further indicated that AtFLS1 contributes to plant defense against Cyst nematode infection, while other AtFLS gene family members do not, consistent with prior reports that these other genes encode little or no enzyme activity. Together, these findings indicate a role of AtMyb12 in promoting the early stages of Cyst nematode infection, while flavonols produced through the action of AtFLS1 are essential for plant defense. On the other hand, a transient induction of AtMYB12 was not observed in galls produced during root-knot nematode infection, but this gene was instead substantially downregulated, starting at the 9 dpi sampling point, as were AtCHS and AtFLS1. In addition, both the AtMYB12- and AtFLS1-deficient lines were more susceptible to infection by this parasite. There was again little evidence for contributions from the other AtFLS gene family members, although an AtFLS5-deficient line appeared to be somewhat more susceptible to infection. Taken together, this study shows that sugar-beet cyst and root-knot nematodes modulate differently the genes involved in flavonol biosynthesis in order to successfully infect host roots and that AtFLS1 may be involved in the plant basal defense response against nematode infection

Expression of Arabidopsis pathogenesis-related genes during nematode infection

P>The expression pattern of pathogenesis-related genes PR-1 to PR-5 was examined in the roots and leaves of Arabidopsis thaliana plants on infection with beet-cyst (Heterodera schachtii) and root-knot (Meloidogyne incognita) nematodes. During H. schachtii parasitism of Arabidopsis, the expression of PR-1, PR-2 and PR-5, which are considered to be markers for salicylic acid (SA)-dependent systemic acquired resistance (SAR), was induced in both roots and leaves of infected plants. In addition, the expression of PR-3 and PR-4, which are used as markers for jasmonic acid (JA)-dependent SAR, was not altered in roots, but in the leaves of H. schachtii-infected plants, the expression PR-3 was induced, whereas the expression of PR-4 was down-regulated. During M. incognita infection of Arabidopsis, the expression of PR-1, PR-2 and PR-5 was highly induced in roots, as was PR-3 to a lesser extent, but the expression of PR-4 was not altered, indicating that infection with M. incognita activated both SA- and JA-dependent SAR in roots. However, all PRgenes examined (PR-1 to PR-5) were down-regulated in the leaves of M. incognita-infected plants, suggesting the suppression of both SA- and JA-dependent SAR. Furthermore, constitutive expression of a single PR in Arabidopsis altered the transcription patterns of other PR genes, and the over-expression of PR-1 reduced successful infection by both H. schachtii and M. incognita, whereas the over-expression of PR-3 reduced host susceptibility to M. incognita but had no effect on H. schachtii parasitism. The results suggest that fundamental differences in the mechanisms of infection by beet-cyst and root-knot nematodes differentially regulate PR protein production and mobilization within susceptible host plants