Project description:Transcriptomes analysis of the petals from a red-flowered white clover mutant (red flowers) and its shade-treated counterpart (white flowers) grown under shaded conditions was performed using high-throughput sequencing. We obtained 121,626,564 and 130,577,944 clean reads in red-flowered mutant and treated counterpart, respectively. Of these clean reads, we respectively gained 125,350 and 99,638 unigene sequencces in two groups. As a result, a total of 157,964 unigenes were obtained with an average length of 728 bp and a median length of 1346 bp. These findings provideed insights into the expression profiles in red-flowered white clover mutant, and deepened our understanding of flower pigmentation in white clower.
Project description:Red clover (Trifolium pratense L.) is one of the most important legume forage species in temperate livestock agriculture. Tetraploid red clover cultivars are generally producing less seed than diploid cultivars. Improving the seed setting potential of tetraploid cultivars is necessary in order to utilize the high forage quality and environmentally sustainable nitrogen fixation of red clover. Two genotypes, one from cv.Tripo with weak seed setting and one from cv.Lasang with strong seed setting, were selected based on data from field experiments for transcriptome analysis of developing flower buds. De novo and reference based analyses of transcriptome assemblies were conducted to study the global transcriptome changes from early to late developmental stages of flower development of the two contrasting red clover genotypes. Transcript profiles, gene ontology enrichment and KEGG pathway analysis indicate that genes related to flower development, pollen pistil interactions, photosynthesis and embryo development are differentially expressed between the Tripo and Lasang genotypes. A significant number of genes related to pollination was overrepresented in Lasang, which might be a reason for its good seed setting ability. The candidate genes detected in this study might be used to develop molecular tools for breeding tetraploid red clover varieties with improved seed yield potentials.
Project description:White clover mosaic virus (WCMV) is a major pathogen of white clover (Trifolium repens L.), with significant effects on yield and persistence. Due to the absence of natural sources of WCMV resistance a transgenic strategy has been employed to produce plants constitutively expressing WCMV replicase gene derivatives, designed to inhibit the propagation of WCMV through an RNA silencing mechanism. A 12,000 feature oligonucleotide microarray has been used to identify global changes in host plant, in addition to virus genome-encoded gene expression associated with WCMV infection in non-transgenic and transgenic WCMV-resistant white clover. Pairwise comparison between the transcriptome of mock-inoculated non-transgenic and WCMV-inoculated transgenic plants provides clear evidence for substantial equivalence between these two genotype/treatments, and demonstrate the efficacy of the transgenic strategy. WCMV- inoculated non-transgenic plants exhibit elevated abundance of many virus-encoded, and host immune response-specific transcripts compared to the transgenic resistant plants or mock-inoculated non-transgenic plants. By contrast, relative to inoculated sensitive plants, the majority of significantly up-regulated genes in mock-inoculated non-transgenic plants or WCMV-inoculated transgenic plants are markers of healthy cellular function. These results, and the occurrence of levels of WCMV-encoded transcripts in inoculated transgenic plants equivalent to those in virus-free plants, confirm the validity of the transgenic RNA silencing approach.<br>
Project description:12plex_medicago_2014_02 - nar nodule vs root transcriptome - which are the genes differentially expressed in alfalfa spontaneous (non rhizobium-infected) nodules vs. control roots? - biological material: aeroponically grown cuttings of a Medicago sativa (alfalfa) accession that produces empty nodules when nitrogen-starved. Samples for transcriptome comparison: isolated NAR nodules (10 days post N-starvation) vs. roots of the same plants (pools of 3 roots).
Project description:Legumes establish endosymbiotic associations with nitrogen-fixing rhizobia, which they host inside root nodules. Here, specific physiological and morphological adaptations, such as the production of oxygen-binding leghemoglobin proteins and the formation of an oxygen diffusion barrier in the nodule periphery, are essential to protect the oxygen-labile bacterial nitrogenase enzyme. The molecular basis of the latter process remains elusive, as the identification of required genes is limited by the epistatic effect of nodule organogenesis over nodule infection and rhizobia accommodation. We overcame this by exploring the phenotypic diversity of Lotus japonicus accessions that uncouple nodule organogenesis from nodule infection when inoculated with a sub-compatible Rhizobium strain. Using comparative transcriptomics, we identified genes with functions associated with oxygen homeostasis and deposition of lipid polyesters on cell walls to be specifically upregulated in infected compared to uninfected nodules. As such hydrophobic modifications on cell walls are pivotal for creating diffusion barriers like the root endodermis, we focused on two Fatty acyl-CoA reductase genes that were specifically activated in the nodule or in the root endodermis. Mutant lines in a Fatty acyl-CoA reductase gene expressed exclusively in the nodule endodermis showed had decreased suberization of this cell layer and increased nodule permeability compared to wild type plants. Oxygen concentrations were significantly increased in the inner cortex of mutant nodules, which correlated with reduced nitrogen fixation rates, and impaired shoot growth. These results provide the first genetic evidence for the formation of the nodule oxygen diffusion barrier, a key adaptation enabling nitrogen fixation in legume nodules.
Project description:Improvement of freezing tolerance of red clover (Trifolium pratense L.) would increase its persistence under cold climate. In this study, we assessed the freezing tolerance and compared the proteome composition of non-acclimated and cold-acclimated plants of two initial cultivars of red clover: Endure (E-TF0) and Christie (C-TF0) and of populations issued from these cultivars after three (TF3) and four (TF4) cycles of phenotypic recurrent selection for superior freezing tolerance. Through this approach, we wanted to identify proteins that are associated with the improvement of freezing tolerance in red clover. Recurrent selection performed indoor is an effective approach to improve the freezing tolerance of red clover. Significant improvement of freezing tolerance by recurrent selection was associated with differential accumulation of a small number of cold-regulated proteins that may play an important role in the determination of the level of freezing tolerance.
Project description:Coevolutionary change requires reciprocal selection between interacting species, i.e., that the partner genotypes that are favored in one species depend on the genetic composition of the interacting species. Coevolutionary genetic variation is manifested as genotype ´ genotype (G ´ G) interactions for fitness from interspecific interactions. Although quantitative genetic approaches have revealed abundant evidence for G ´ G interactions in symbioses, the molecular basis of this variation remains unclear. Here we study the molecular basis of G ´ G interactions in a model legume-rhizobium mutualism using gene expression microarrays. We find that, like quantitative traits such as fitness, variation in the symbiotic transcriptome may be partitioned into additive and interactive genetic components. Our results suggest that plant genetic variation is the largest influence on nodule gene expression, and that plant genotype and the plant genotype ´ rhizobium genotype interaction determine global shifts in rhizobium gene expression that in turn feedback to influence plant fitness benefits. Moreover, the transcriptomic variation we uncover implicates regulatory changes in both species as drivers of symbiotic gene expression variation. Our study is the first to partition genetic variation in a symbiotic transcriptome, and illuminates potential molecular routes of coevolutionary change. We assayed gene expression using three biological replicates for each plant genotype × rhizobium genotype combination (4 combinations) for a total of 12 chips.