TU16032 Optimizing 10x technology (BO-56-201-001, BO-50-003-001, BO-25.08-003-001)

Crops and other plants live in close association with millions of microorganisms, which constitute the microbiome of their rhizosphere and the surrounding soil. Soil and plant microbiomes contribute substantially to the growth and development of the plant, as well as to its resilience against abiotic stress, pests and diseases. Within current agricultural practice, there is an urgent need for novel green solutions based on natural (micro)organisms and compounds to combat plant pathogens, increase plant resilience against drought and achieve sustainable cultivation with minimal use of water and nutrients. The plant microbiome offers potential solutions for these challenges.

Recent research suggests that specific microbial specialized metabolites—particularly terpenes, nonribosomal peptides and polyketides—play a key role in microbe-mediated beneficial effects on plant growth- and health. Innovative computational techniques now allow rapid identification and classification of biosynthetic gene clusters (BGCs) that encode the enzymatic pathways underlying the production of these compounds. The NWO Groen project ‘Harnessing the soil microbiome for improved stress tolerance in crop plants’, coordinated by Dr. Medema at Wageningen University, will use an innovative metagenome-wide association study to systematically connect multiple desirable and quantitatively measurable crop phenotypes across a large number of soil samples to the BGCs and microbial metabolites responsible for these traits. The obtained results will allow us to develop new products (microbes and metabolites) to combat various types of biotic (fungal pathogens) and abiotic (soil drought) stress.

A key technological bottleneck in this approach is that it is often difficult to assemble full-length BGCs in metagenomes from complex microbiomes, such as the rhizosphere. The entire BGC sequence is required in order to fully assess chemical novelty of its molecular product, and to allow heterologous expression or refactoring through DNA synthesis. A promising technology that could vastly improve metagenomic assembly in a cost-effective manner is 10x Genomics. At the beginning of 2016, the company launched their Chromium platform. The core technology is a Gel bead inside Emulsion (GEM) and uses droplet microfluidic technology prior to standard (Illumina) sequencing to generate “linked reads” that can be used to scaffold or bin metagenomic contigs. Examples of success currently focus on whole-genome sequencing, in particular the diploid human genome (www.10xgenomics.com). We propose to early adopt 10x Genomics technology on metagenomes and to develop methods for improving metagenome assembly customized for this technology. To do so, we here propose research to optimise the technology (in terms of DNA fragment length, coverage, assembly) for use on metagenomic data, so that it can be effectively used to discover crop protection agents from natural products. Collectively, this research will strongly support the NWO Groen project in gaining a better understanding of the interactions between plants, pathogens and beneficial microbiota, leading to new products to combat crop diseases and more sustainable crop cultivation.