Symbiosis-dependent microbiome patterning in Lotus japonicus roots

对豆科植物百脉根中的许多微生物组进行测序，为了解根际内生菌的复杂性以及与宿主根的相关性提供了很好的见解。然而，我们对时空定殖模式以及这些细胞间定殖细菌和真菌之间的任何串扰的了解很少，因为这些信息无法从全局测序数据中检索到。此外，几乎所有的豆科植物的遗传研究，评估其功能的控制共生微生物，如根瘤菌和丛枝菌根真菌（AMF）的殖民宿主基因已通过应用单关联设置进行。然而，在自然土壤中，情况是根本不同的，豆类根部同时被多种微生物定殖。因此，该提议旨在通过解决以下方面来增加实验装置的复杂性：1. 评估一系列共生突变体在自然土壤中的表现。基于我们的试点实验，我们将筛选一系列Lotus突变体，以了解它们绕过目前被认为是RNS不可或缺的调控网络的能力。将鉴定定殖这些突变体的菌株。2. 大规模测试的单协会最初使用70个菌株的细菌核心收集，从莲藕产生。最有效的殖民者将被组合成一个最小的合成社区（最小的SynCom）。 在最小SynCom内产生标记菌株。4. 分析了这些标记菌株在寄主根内的分布模式，并初步分析了控制它们的可能寄主因素。

Sequencing a number of microbiomes from the legume Lotus japonicus provided great insights into the complexity of endophytes present in the rhizosphere and associated with the host root. However, our knowledge on spatio-temporal colonization patterns and any cross-talk in between these intercellularly colonizing bacteria and fungi is sparse as these information cannot be retrieved from global sequencing data. Furthermore, almost all genetic studies in legumes that assessed host genes for their function in controlling colonization by symbiotic microbes such as rhizobia and arbuscular mycorrhiza fungi (AMF) have been conducted by applying mono-association setups. However, the situation is fundamentally different in natural soils, where legume roots are colonized by multiple microbes simultaneously. Therefore, this proposal aims to increase complexity in the experimental setup by addressing the following aspects:1. Assessing a range of symbiosis mutants for their performance in natural soils. Based on our pilot experiments we will screen a collection of Lotus mutants for their ability to bypass regulatory networks that are currently considered to be indispensable for RNS. Strains colonizing these mutants will be identified. 2. Large-scale testing of mono-associations initially using 70 isolates from a bacterial core collection that was generated from Lotus japonicus roots. The most effective colonizers will be combined to a minimal synthetic community (minimal SynCom).3. Generation of marker strains within the minimal SynCom.4. Analysing distribution patterns of these marker strains within the host root and initial analysis of possible host factors controlling them.