Project description:Identification of microRNA targets in M. truncatula roots Degradome sequencing of mycorrhizal and non-mycorrhiza M. truncatula roots
Project description:Identification of microRNA expressed in M. truncatula roots Small RNA sequencing of mycorrhizal and non-mycorrhiza M. truncatula roots
Project description:Composite plants consisting of a wild-type shoot and a transgenic root are frequently used for functional genomics in legume research. Although transformation of roots using Agrobacterium rhizogenes leads to morphologically normal roots, the question arises as to whether such roots interact with arbuscular mycorrhizal (AM) fungi in the same way as wild-type roots. To address this question, roots transformed with a vector containing the fluorescence marker DsRed were used to analyse AM in terms of mycorrhization rate, morphology of fungal and plant subcellular structures, as well as transcript and secondary metabolite accumulations. Mycorrhization rate, appearance, and developmental stages of arbuscules were identical in both types of roots. Using Mt16kOLI1Plus microarrays, transcript profiling of mycorrhizal roots showed that 222 and 73 genes exhibited at least a 2-fold induction and less than half of the expression, respectively, most of them described as AM regulated in the same direction in wild-type roots. To verify this, typical AM marker genes were analysed by quantitative reverse transcription-PCR and revealed equal transcript accumulation in transgenic and wild-type roots. Regarding secondary metabolites, several isoflavonoids and apocarotenoids, all known to accumulate in mycorrhizal wild-type roots, have been found to be up-regulated in mycorrhizal in comparison with non-mycorrhizal transgenic roots. This set of data revealed a substantial similarity in mycorrhization of transgenic and wild-type roots of Medicago truncatula, validating the use of composite plants for studying AM-related effects.
Project description:BACKGROUND: Most vascular flowering plants have the capacity to form symbiotic associations with arbuscular mycorrhizal (AM) fungi. The symbiosis develops in the roots where AM fungi colonize the root cortex and form arbuscules within the cortical cells. Arbuscules are enveloped in a novel plant membrane and their establishment requires the coordinated cellular activities of both symbiotic partners. The arbuscule-cortical cell interface is the primary functional interface of the symbiosis and is of central importance in nutrient exchange. To determine the molecular events the underlie arbuscule development and function, it is first necessary to identify genes that may play a role in this process. Toward this goal we used the Affymetrix GeneChip Medicago Genome Array to document the M. truncatula transcript profiles associated with AM symbiosis, and then developed laser microdissection (LM) of M. truncatula root cortical cells to enable analyses of gene expression in individual cell types by RT-PCR. RESULTS: This approach led to the identification of novel M. truncatula and G. intraradices genes expressed in colonized cortical cells and in arbuscules. Within the arbuscule, expression of genes associated with the urea cycle, amino acid biosynthesis and cellular autophagy was detected. Analysis of gene expression in the colonized cortical cell revealed up-regulation of a lysine motif (LysM)-receptor like kinase, members of the GRAS transcription factor family and a symbiosis-specific ammonium transporter that is a likely candidate for mediating ammonium transport in the AM symbiosis. CONCLUSION: Transcript profiling using the Affymetrix GeneChip Medicago Genome Array provided new insights into gene expression in M. truncatula roots during AM symbiosis and revealed the existence of several G. intraradices genes on the M. truncatula GeneChip. A laser microdissection protocol that incorporates low-melting temperature Steedman's wax, was developed to enable laser microdissection of M. truncatula root cortical cells. LM coupled with RT-PCR provided spatial gene expression information for both symbionts and expanded current information available for gene expression in cortical cells containing arbuscules.
Project description:Arbuscular mycorrhizal (AM) fungi establish symbiosis and improve the lead (Pb) tolerance of host plants. The AM plants accumulate more Pb in roots than their non-mycorrhizal counterparts. However, the direct and long-term impact of AM fungi on plant Pb uptake has been rarely reported. In this study, AM fungus (Rhizophagus irregularis) colonized and non-colonized roots of Medicago truncatula were separated by a split-root system, and their differences in responding to Pb application were compared. The shoot biomass accumulation and transpiration were increased after R. irregularis inoculation, whereas the biomass of both colonized and non-colonized roots was decreased. Lead application in the non-colonized root compartment increased the R. irregularis colonization rate and up-regulated the relative expressions of MtPT4 and MtBCP1 in the colonized root compartments. Rhizophagus irregularis inoculation increased Pb uptake in both colonized and non-colonized roots, and R. irregularis transferred Pb to the colonized root segment. The Pb transferred through the colonized root segment had low mobility and might be sequestrated and compartmented in the root by R. irregularis. The Pb uptake of roots might follow water flow, which is facilitated by MtPIP2. The quantification of Pb transfer via the mycorrhizal pathway and the involvement of MtPIP2 deserve further study.
Project description:affy_med_2011_09: In natural ecosystems most vascular plants develop symbiosis with arbuscular mycorrhizal (AM) fungi which help them acquire nutrients such as phosphorus (P) and nitrogen (N). P has long been known to control AM symbiosis which takes place only when P is limiting. For N, however, its role in controlling mycorrhization is less clear. We have chosen the model plant Medicago truncatula to analyze the impact of P limitation and both P and N limitation on Medicago root transcriptome in response to the AM fungus Rhizophagus irregularis (formerly Glomus intraradices (BEG141)). These analyses may help us uncover signaling events involved in the interaction between these symbionts as well as genes encoding transporters potentially important for nutrient exchanges in these conditions. --We will compare the root transcriptome of Medicago truncatula plants inoculated with Rhizophagus irregularis to that of non-inoculated plants grown under P limitation (or both P and N limitation) after 4 weeks of culture
Project description:The nomenclatural type material of Rhizophagus intraradices (basionym Glomus intraradices) was originally described from a trap pot culture established with root fragments, subcultures of which later became registered in the INVAM culture collection as FL 208. Subcultures of FL 208 (designated as strain ATT 4) and a new strain, independently isolated from the type location (ATT 1102), were established as both pot cultures with soil-like substrate and in vitro root organ culture. Long-term sampling of these cultures shows spores of the species to have considerable morphological plasticity, not described in the original description. Size, shape and other features of the spores were much more variable than indicated in the protologue. Phylogenetic analyses confirmed earlier published evidence that sequences from all R. intraradices cultures formed a monophyletic clade, well separated from, and not representing a sister clade to, R. irregularis. Moreover, new phylogenetic analyses show that Rhizoglomus venetianum and R. irregularis are synonymous. The morphological characters used to separate these species exemplify the difficulties in species recognition due to the high phenotypic plasticity in the genus Rhizophagus. Rhizophagus intraradices is morphologically re-described, an epitype is designated from a single-spore isolate derived from ATT 4, and R. venetianum is synonymised with R. irregularis.