Transcriptomic analysis of Medicago truncatula calli with MtWOX9-1 overexpression
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ABSTRACT:
ORGANISM(S): Medicago truncatula
PROVIDER: GSE201314 | GEO | 2022/04/25
REPOSITORIES: GEO
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Project description:Somatic embryogenesis (SE), a morphogenic process that takes advantage of the regenerative potential of plants to replicate whole plants starting from somatic explants, can be a source of variation with potential applications in plant breeding. It is one of the most suitable tools to apply functional genomics studies and genetic improvement in plants. However, beyond a few pioneering works mainly focused on model plants, the molecular characterization of SE mechanisms is still elusive, especially for woody species. In grapevine, this process is affected by many factors such as explant type, culture conditions and, most importantly, genotype. Many cultivars, in fact, have shown recalcitrance to tissue culture and transformation, and very low SE competence. Thus, the understanding of SE competence behind the regenerative aptitude is fundamental to the widespread application of the so-called “next-generation breeding techniques”, such as cisgenesis and genome editing, in grapevine. Here, we explored genetic and epigenetic features of the SE process in grapevine by investigating the behavior of two genotypes showing opposite SE competence. Embryogenic tissues were induced from immature stamens excised from field-collected flower clusters of Sangiovese (highly competent for embryogenesis) and Cabernet Sauvignon (poorly competent for embryogenesis). A multilayered approach was used to profile mRNA, smallRNAs and methylated DNA with high-throughput sequencing technologies in the initial explants, on undifferentiated calli induced after 40 days of culture, and in embryogenic and non-embryogenic calli after 3 months of culture. A comprehensive comparison of transcriptomes of the different types of calli with the grapevine gene expression atlas revealed that, in grapevine, the dedifferentiation to the callus formation during the embryogenesis process occurs via a ‘berry’ developmental pathway. This is dissimilar from that shown in Arabidopsis, in which dedifferentiated calli are more similar to the tip of a root meristem. Interestingly, a Gene Ontology (GO) analysis revealed that secondary metabolism and gene expression regulation/epigenetics are the enriched functional categories of genes differentially down- and up-regulated in embryogenic vs non-embryogenic calli, respectively. These results prompted us to define the epigenetic landscape dynamics during SE in grapevine, revealing a significant increase in DNA methylation, especially in intergenic regions, in the embryogenic calli tissues. Finally, we proposed potential key regulators of SE in different genotypes that could represent putative targets of next-generation breeding techniques in grapevine.
2021-07-07 | GSE133761 | GEO
Project description:Somatic embryogenesis (SE), a morphogenic process that takes advantage of the regenerative potential of plants to replicate whole plants starting from somatic explants, can be a source of variation with potential applications in plant breeding. It is one of the most suitable tools to apply functional genomics studies and genetic improvement in plants. However, beyond a few pioneering works mainly focused on model plants, the molecular characterization of SE mechanisms is still elusive, especially for woody species. In grapevine, this process is affected by many factors such as explant type, culture conditions and, most importantly, genotype. Many cultivars, in fact, have shown recalcitrance to tissue culture and transformation, and very low SE competence. Thus, the understanding of SE competence behind the regenerative aptitude is fundamental to the widespread application of the so-called “next-generation breeding techniques”, such as cisgenesis and genome editing, in grapevine. Here, we explored genetic and epigenetic features of the SE process in grapevine by investigating the behavior of two genotypes showing opposite SE competence. Embryogenic tissues were induced from immature stamens excised from field-collected flower clusters of Sangiovese (highly competent for embryogenesis) and Cabernet Sauvignon (poorly competent for embryogenesis). A multilayered approach was used to profile mRNA, smallRNAs and methylated DNA with high-throughput sequencing technologies in the initial explants, on undifferentiated calli induced after 40 days of culture, and in embryogenic and non-embryogenic calli after 3 months of culture. A comprehensive comparison of transcriptomes of the different types of calli with the grapevine gene expression atlas revealed that, in grapevine, the dedifferentiation to the callus formation during the embryogenesis process occurs via a ‘berry’ developmental pathway. This is dissimilar from that shown in Arabidopsis, in which dedifferentiated calli are more similar to the tip of a root meristem. Interestingly, a Gene Ontology (GO) analysis revealed that secondary metabolism and gene expression regulation/epigenetics are the enriched functional categories of genes differentially down- and up-regulated in embryogenic vs non-embryogenic calli, respectively. These results prompted us to define the epigenetic landscape dynamics during SE in grapevine, revealing a significant increase in DNA methylation, especially in intergenic regions, in the embryogenic calli tissues. Finally, we proposed potential key regulators of SE in different genotypes that could represent putative targets of next-generation breeding techniques in grapevine.
2021-12-28 | GSE152520 | GEO
Project description:Somatic embryogenesis (SE), a morphogenic process that takes advantage of the regenerative potential of plants to replicate whole plants starting from somatic explants, can be a source of variation with potential applications in plant breeding. It is one of the most suitable tools to apply functional genomics studies and genetic improvement in plants. However, beyond a few pioneering works mainly focused on model plants, the molecular characterization of SE mechanisms is still elusive, especially for woody species. In grapevine, this process is affected by many factors such as explant type, culture conditions and, most importantly, genotype. Many cultivars, in fact, have shown recalcitrance to tissue culture and transformation, and very low SE competence. Thus, the understanding of SE competence behind the regenerative aptitude is fundamental to the widespread application of the so-called “next-generation breeding techniques”, such as cisgenesis and genome editing, in grapevine. Here, we explored genetic and epigenetic features of the SE process in grapevine by investigating the behavior of two genotypes showing opposite SE competence. Embryogenic tissues were induced from immature stamens excised from field-collected flower clusters of Sangiovese (highly competent for embryogenesis) and Cabernet Sauvignon (poorly competent for embryogenesis). A multilayered approach was used to profile mRNA, smallRNAs and methylated DNA with high-throughput sequencing technologies in the initial explants, on undifferentiated calli induced after 40 days of culture, and in embryogenic and non-embryogenic calli after 3 months of culture. A comprehensive comparison of transcriptomes of the different types of calli with the grapevine gene expression atlas revealed that, in grapevine, the dedifferentiation to the callus formation during the embryogenesis process occurs via a ‘berry’ developmental pathway. This is dissimilar from that shown in Arabidopsis, in which dedifferentiated calli are more similar to the tip of a root meristem. Interestingly, a Gene Ontology (GO) analysis revealed that secondary metabolism and gene expression regulation/epigenetics are the enriched functional categories of genes differentially down- and up-regulated in embryogenic vs non-embryogenic calli, respectively. These results prompted us to define the epigenetic landscape dynamics during SE in grapevine, revealing a significant increase in DNA methylation, especially in intergenic regions, in the embryogenic calli tissues. Finally, we proposed potential key regulators of SE in different genotypes that could represent putative targets of next-generation breeding techniques in grapevine.
2021-12-28 | GSE152337 | GEO
Project description:Carotenoids have been demonstrated to be indispensable plant secondary metabolites that are involved in photosynthesis, antioxidation, and phytohormone biosynthesis. Carotenoids are likely involved in other biological functions that have yet to be discovered. In this study, we utilized genomic expression investigation to gain a deep insight into the carotenoid-related biological processes in the citrus calli overexpressing CrtB. Abortive ovule embryogenic calli from four citrus genotypes were used in this study. They were derived from Star Ruby grapefruit (C. paradise Macf.), Marsh grapefruit (C. paradise Macf.), and Sunburst mandarin [Citrus reticulata Blanco M-CM-^W (C. paradisi Macf. M-CM-^W C. reticulata)], designated as RB, M, and SBT, respectively. Engineered cell models (ECMs) were established by over-expressing 35S::CrtB (tpM-bM-^@M-^SrbcSM-bM-^@M-^SCrtB) [CrtB protein, phytoene synthase from Erwinia herbicola (now known as Pantoea agglomerans), containing a Pea rbcS transit peptide] in citrus embryogenic calli. Twenty-day-old calli were harvested and used for RNA extraction and hybridization on Affymetrix microarrays.
2014-09-23 | E-GEOD-61633 | biostudies-arrayexpress
Project description:Understanding the mechanisms that endow a somatic cell with the ability to differentiate into a somatic embryo, which could result in numerous biotechnological applications, is still a challenge. The objective of this work was to identify some of the molecular and physiological mechanisms responsible for the acquisition of embryogenic competence during somatic embryogenesis in Carica papaya L. We performed a broad characterization of embryogenic (ECs) and nonembryogenic calli (NECs) of C. papaya using global and mitochondrial proteomic approaches, histomorphology, histochemistry, respiratory activity, and endogenous hormonal and hydrogen peroxide contents. ECs and NECs presented remarkable differences in anatomical and histochemical characteristics. ECs showed greater reactivity for the presence of proteins and neutral polysaccharides. Our results demonstrate the role of mitochondrial metabolism in the embryogenic competence of C. papaya calli. Greater participation of alternative oxidase (AOX) enzymes in respiration, as well as stronger accumulation of mitochondrial stress response proteins, was observed in ECs. In addition, ECs showed a greater abundance of proteins related to oxidative phosphorylation and higher total respiration (TR). Auxin-responsive Gretchen Hagen 3 (GH3) family proteins may play an important role in decreasing the contents of free 2,4-dichlorophenoxyacetic acid (2,4-D) in ECs. The accumulation of stress response proteins among total proteins was observed in ECs. ECs also showed higher endogenous hydrogen peroxide (H2O2) contents. H2O2 is a promising molecule for further investigation in differentiation protocols for C. papaya somatic embryos.
2022-06-28 | PXD026311 | Pride
Project description:Several factors influence the culture conditions and somatic embryogenesis responses, such as the role of plant growth regulators (PGRs) during the establishment of in vitro cultures. For instance, auxin is required for callus induction and the acquisition of embryogenic capacity in different species, but the removal of auxin is needed for the further differentiation of somatic embryos. Thus, we aimed to evaluate the effects of residual 2,4-dichlorophenoxyacetic acid (2,4-D) on the maturation of sugarcane somatic embryos. First, embryogenic calli were separated into two groups: the PGR-free group was cultured without 2,4-D and b) the control group was maintained on proliferation culture medium, which contains 2,4-D. After 21 days of culture in the dark, both groups were transferred to maturation culture medium under light. Our results showed that calli grown in PGR-free culture medium yielded more somatic embryos and exhibited a higher accumulation of protein and starch reserves. A proteomic analysis showed a decrease in the abundance of abscisic acid (ABA)-induced proteins in the control group. The levels of residual 2,4-D and endogenous 1-aminocyclopropane-1-carboxylic acid (ACC), an ethylene precursor, were higher in the control group than in the PGR-free group. Conversely, the level of ABA was higher in the PGR-free group than in the control group. A disruption in the ABA and ethylene levels might be responsible for the observed delay in the accumulation of storage reserves in the embryogenic calli of the control group, which might have affected the development of somatic embryos. Our results also showed that the efficient development of sugarcane somatic embryos appears to be preceded by an efficient accumulation of storage reserves in calli, which is related to hormone homeostasis.
2021-06-01 | PXD021987 | Pride
Project description:Agrobacterium tumefaciens-mediated genetic transformation has been routinely used in rice for more than a decade. However, the transformation efficiency of the indica rice variety is still unsatisfactory and much lower than that of japonica cultivars. Further improvement on the transformation efficiency lies in the genetic manipulation of the plant itself, which requires a better understanding of the underlying process accounting for the susceptibility of plant cells to Agrobacterium infection as well as the identification of plant genes involved in the transformation process. In order to investigate the related genes affecting the transformation efficiency of embryogenic calli of different rice cultivars, we used Affymetrix GeneChipM-BM-. Rice Genome Array to measure the global gene expression profiling just before transformation and at four different time points after transformation (1 h, 6 h, 12 h, 24 h) in both japonica rice cultivar Nipponbare and indica rice cultivar Zhenshan 97. The mature embryo-derived embryogenic calli of Nipponbare (Nip) and Zhenshan 97 (ZS) were infected by Agrobacterium. Calli of Nip and ZS were sampled just before infection (0 h) and 1h, 6h, 12 h and 24h after infection, respectively. Three independent biological replications for each time point of the two varieties were used. To avoid the influence of polymorphisms between the probe sequence on the array and the genomes of the varieties used, we used a genomic DNA (gDNA)-based probe-selection strategy based on the hybridization efficiency of gDNA from Nip and ZS with the PM oligonucleotide probes on the rice array. The genomic DNA of Nip and ZS were extracted and hybridized to the Affymetrix Rice Genome Arrays. Three biological replications per cultivar were performed.
2012-01-01 | E-GEOD-32426 | biostudies-arrayexpress
Project description:Somatic embryogenesis (SE) is a complex stress related process regulated by numerous biological factors. SE is mainly applicable to mass propagation and genetic improvement of plants through gene transfer technology and mutation breeding. In banana, SE is highly genome dependent as the efficiency varies with cultivars. To understand molecular mechanism of SE, proteomics approach was carried out to identify genes responsible for embryogenic calli (EC) induction, regeneration and germination of somatic embryos (se) in cv. Rasthali (AAB). In total, 70 spots were differentially expressed in various developmental stages of SE. Of which, 16 were uniquely expressed and 17 were highly abundant in EC than nonembryogenic calli and explant and four spots were also uniquely expressed in germinating se. Functional annotation of identified proteins revealed that calcium signaling along with stress and endogenous hormones related proteins played a vital role in EC induction and germination of se. Thus based on the outcome, callus induction media was modified and tested in five cultivars. In cv. Grand Naine (AAA), increased concentration of 3- IAA and tryptophan recorded highest EC induction of 24.28% while Red Banana with similar genome showed 18.96% in kinetin supplemented media. Similarly, in cultivars Monthan and Karpuravalli with ABB genome showed maximum EC induction in tryptophan supplemented media (8.54%) and CaCl2 enriched media (17.34%) respectively. In cv. Neypoovan (AB), higher concentration of tryptophan induced more EC. These results illustrated that EC formation is genome as well as cultivar dependent. Simultaneously, germination media was modified to induce proteins responsible for germination. In cv. Rasthali, media supplemented with 10 mM CaCl2 showed maximum increase in germination (51.79%) over control. Thus present study revealed that media modification based on proteomic studies can induce SE in recalcitrant cultivars and also enhance germination in cultivars amenable for SE.
2019-03-19 | PXD011392 | Pride
Project description:To identify transcription factor (TF) genes exhibiting preferential expression during SE, expression profiles of 1,880 TFs were compared in two genotypes with largely different capacities for SE induction in IZE culture on auxin medium, namely the highly embryonic Col-0 accession and the tanmei mutant unable to form somatic embryos. Our study revealed 729 TFs whose expression changes during the 10-day incubation period of SE; 141 TFs displayed distinct differences in expression patterns in embryogenic versus non-embryogenic cultures. This study provides comprehensive data focused on the expression of TF genes during SE and provides guidelines for further research on functional genomics of SE. The experiment was designed to monitor the expression of 1,880 TF genes at three distinctive stages of IZE-derived embryogenic culture, which refer to: (i) freshly isolated explants competent to undergo embryogenic transition (0 d), (ii) explant tissue subjected to SE induction (5 d), and (iii) the advanced phase of embryogenesis related to somatic embryo formation (10 d).
2013-07-31 | E-GEOD-45697 | biostudies-arrayexpress
Project description:Transcriptomic analysis of Medicago truncatula calli with MtWOX9-1 overexpression
| PRJNA830848 | ENA