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Negative interactions between the organelle and nuclear genomes (cytonuclear incompatibility) are thought to be among the early genetic incompatibilities to arise during speciation. While there are now several good examples of cytonuclear incompatibility leading to reproductive isolation within and between closely related species, there are still many unanswered questions regarding the evolutionary dynamics of cytonuclear incompatibility. Using the herbaceous species Campanulastrum americanum, I examine how patterns of organelle inheritance and rates of organelle genome evolution may both facilitate and constrain the evolution of cytonuclear incompatibility and its ability to drive the early stages of speciation. [mehr]

Second generation sequencing has caused major breakthroughs in the use of archival DNA, and in the use of herbarium specimens in particular. Whereas this enables testing of historical biological hypotheses, concerns remained about accuracy of herbarium sequence data and the possibility of post-mortem damage. Using a panel of angiosperm trees we compared fresh and historic samples of the same individuals and concluded that such damage is negligible, and that specimen age per se does not predict sequencing success. 2nd generation sequencing retrieves herbarium plastomes surprisingly well, which opens up possibilities for further taxonomic and time sampling. One such clades is Pelargonium (Geraniaceae) which is well-known for its horticultural importance as well as its elevated levels of (organellar) genomic evolution, or genome instability. Whereas some family members have lost Inverted Repeats alltogether, the hybrid P x hortorum is usually cited as having the largest Inverted Repeats known in angiosperms. However, to what extend this is a natural phenomenon or a ‘breeding artefact’ remains to be ascertained. In addition, it is in this Pelargonium clade that both bi-parental inheritance and cytonuclear incongruence occurs. In the context of increased taxonomic sampling of plastomes around the parent species of P x hortorum, be it from herbarium or fresh material, we hope to elucidate this and other questions further. [mehr]

Antibodies are a popular tool used in plant cell biology research. They can be either custom made or purchased from a commercial supplier. In either case their production is a complex process, consisting of three very important components which has to be carefully considered. These are: Antigen-Animal-Testing. Which source of antigen is most optimal for your project: peptide, recombinant protein or a native protein isolated from tissue? Which animal species to choose? Are certain species making better antibodies compare to others? Do I have any controls to validate produced antibody? What controls should be used? What to do if my antibody is not giving any signal in a western blot? [mehr]

A RecG-like DNA helicase is involved in the surveillance of recombination and segregation of plant organellar genomes Clémentine Wallet, Monique Le Ret, André Dietrich and José Gualberto The plasticity of plant mitochondrial genomes is due to recombination processes that modulate their structure. Several factors have already been identified involved in organellar recombination or in recombination-dependent repair processes. But how the segregation of the alternative mitotypes generated by recombination is controlled, is still not understood. An Arabidopsis gene (RECG1) codes for a homologue of bacterial DNA helicase RecG. In bacteria RecG has multiple roles in DNA repair, the control of stoichiometric genome replication and the suppression of ectopic recombination. The plant RECG1 is dually targeted to mitochondria and plastids and can complement bacterial recG deficient strains for repair and replication control. Arabidopsis recG1 mutants have increased ectopic recombination between intermediate size repeat (IR) in mitochondria, and are deficient in repair of double strand breaks induced by a genotoxic stress. In addition we found that RECG1 has roles in the segregation of mtDNA. In a recG1 line an alternative mtDNA sequence generated by recombination is stably maintained as an independent replisome. Reintegration of the wild type RECG1 allele leads to the segregation of the alternative versions of mtDNA, with individual plants inheriting different mtDNA versions. The precise characterization of the recombination steps involved in this sorting process allowed us to build a model for how it is controlled by RECG1. [mehr]

The translation of chloroplast mRNAs has long been known to be subject to regulation by developmental, environmental and physiological cues. However, progress in recognizing examples of translational regulation, identifying translational regulators, and dissecting mechanisms of translational modulation has been limited by the assays that have been available to monitor ribosome behavior in vivo: the traditional assays (pulse-labeling, polysome, and reporter gene approaches) are labor intensive, have limited sensitivity and/or resolution, and are not suited to genome-wide explorations. We are using genome-wide ribosome profiling methods that provide a quantitative and high resolution readout of ribosome positions in vivo to (i) identify nucleus-encoded proteins that are required for the translation of specific chloroplast mRNAs; (ii) analyze the impact of various light regimes on chloroplast ribosome behavior; (iii) describe the translational dynamics of organellar mRNAs during the differentiation of photosynthetic leaf cells; and (iv) elucidate mechanisms involved in the co-translational targeting of proteins to the chloroplast thylakoid membrane. Examples of insights obtained in each of these areas will be discussed. [mehr]