Abstract
BackgroundThorough understanding of complex model systems requires the characterisation of processes in different cell types of an organism. This can be achieved with high-throughput spatial transcriptomics at a large scale. However, for plant model systems this is still challenging as suitable transcriptomics methods are sparsely available. Here we present GaST-seq (Grid-assisted, Spatial Transcriptome sequencing), an easy to adopt, micro-scale spatial-transcriptomics workflow that allows to study expression profiles across small areas of plant tissue at a fraction of the cost of existing sequencing-based methods.ResultsWe compare the GaST-seq method with widely used library preparation methods (Illumina TruSeq). In spatial experiments we show that the GaST-seq method is sensitive enough to identify expression differences across a plant organ. We further assess the spatial transcriptome response of Arabidopsis thaliana leaves exposed to the bacterial molecule flagellin-22, and show that with eukaryotic (Albugo laibachii) infection both host and pathogen spatial transcriptomes are obtained.ConclusionWe show that our method can be used to identify known, rapidly flagellin-22 elicited genes, plant immune response pathways to bacterial attack and spatial expression patterns of genes associated with these pathways.
Highlights
Thorough understanding of complex model systems requires the characterisation of processes in different cell types of an organism
In additional pilot experiments using Arabidopsis thaliana and the oomycete pathogen Albugo laibachii we show that the dual dataset of both host and pathogen, are captured, and spatial expression profiles can be constructed which could be used to study the biotic interactions in infections
Does leaf dissection induce wounding response gene expression profiles? Physical wounding of plants is known to induce wounding related gene expression [19]. This is an important point to consider as the GaST-seq workflow dissects tissue into ~ 1 mm2 squares followed by immediate snap freezing on dry ice
Summary
Thorough understanding of complex model systems requires the characterisation of processes in different cell types of an organism This can be achieved with high-throughput spatial transcriptomics at a large scale. A thorough understanding of these complex systems requires the ability to dissect and characterise processes in the different organs and cell types This is challenging, though recently multiomics single-cell studies have been flourishing [2], but high-throughput, high-resolution methodologies that assess molecular conditions with spatial resolution are sparsely available [3,4,5,6]. GaST-seq is based on three consecutive steps: (1) rapid, mechanical sample dissection of small e.g. 1 mm leaf areas (2) a high-throughput method for high quality mRNA extraction of difficult to lyse plant tissues and (3) generation sequencing (NGS) library construction and sequencing (Fig. 1)
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