SPO11 dependent initiation of meiotic double strand breaks in Arabidopsis thaliana

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The meiotic recombination process depends in nearly all eukaryotes on the initiation of double strand breaks (DSBs) by SPO11, a meiosis specific transesterase. In animals and fungi a single SPO11 gene is sufficient whereas in plants two of them, AtSPO11-1 and 2, are essential for the initiation of meiotic DSBs. Both SPO11 proteins are necessary in a functional form otherwise no DSB induction is performed in Arabidopsis and the plants are nearly sterile due to random chromosome segregation during meiosis. Therefore, we want to investigate whether the functions of these two SPO11 genes in Arabidopsis thaliana are species- and/or sequence-specific or if they can be complemented by Arabidopsis SPO11 genes in which parts of the sequences are interchanged between both genes (swap DNAs). Furthermore, we will use the orthologous SPO11-1 and -2 genes from land plants (Carica papaya and Physcomitrella patens) and cDNAs from lesser related organisms (Chlamydomonas reinhardtii and Mus musculus) for heterologous complementation of SPO11 mutants from Arabidopsis. Using heterologous genes or cDNAs and truncated or partially swapped DNAs of Arabidopsis for the complementation of SPO11-1 and 2 mutants, we should be able to answer the following questions. Is a specific sequence of the respective SPO11 protein necessary for proper initiation of meiotic DSBs and which sequence region is defining the difference between SPO11-1 and 2?