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Artificial micro RNAs (ami RNAs) are used for selective gene silencing in plants. However, current methods to produce ami RNA constructs for silencing transcripts in monocot species are not suitable for simple, cost-effective and large-scale synthesis. Here, a series of expression vectors based on Oryza sativa MIR390 ( Os MIR390) precursor was developed for high-throughput cloning and high expression of ami RNAs in monocots. Four different ami RNA sequences designed to target specifically endogenous genes and expressed from Os MIR390-based vectors were validated in transgenic Brachypodium distachyon plants. Surprisingly, ami RNAs accumulated to higher levels and were processed more accurately when expressed from chimeric Os MIR390-based precursors that include distal stem-loop sequences from Arabidopsis thaliana MIR390a (At MIR390a). In all cases, transgenic plants displayed the predicted phenotypes induced by target gene repression, and accumulated high levels of ami RNAs and low levels of the corresponding target transcripts. Genome-wide transcriptome profiling combined with 5′- RLM- RACE analysis in transgenic plants confirmed that ami RNAs were highly specific. [ABSTRACT FROM AUTHOR]

Plant cells communicate information for the regulation of development and responses to external stresses. A key form of this communication is transcriptional regulation, accomplished via complex gene networks operating both locally and systemically. To fully understand how genes are regulated across plant tissues and organs, high resolution, multi-dimensional spatial transcriptional data must be acquired and placed within a cellular and organismal context. Spatial transcriptomics (ST) typically provides a two-dimensional spatial analysis of gene expression of tissue sections that can be stacked to render three-dimensional data. For example, X-ray and light-sheet microscopy provide sub-micron scale volumetric imaging of cellular morphology of tissues, organs, or potentially entire organisms. Linking these technologies could substantially advance transcriptomics in plant biology and other fields. Here, we review advances in ST and 3D microscopy approaches and describe how these technologies could be combined to provide high resolution, spatially organized plant tissue transcript mapping., (© American Society of Plant Biologists 2021. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

An increase of cytosolic Ca(2+) is generated by diverse physiological stimuli and stresses, including pathogen attack. Plants have evolved two branches of the immune system to defend against pathogen infections. The primary innate immune response is triggered by the detection of evolutionarily conserved pathogen-associated molecular pattern (PAMP), which is called PAMP-triggered immunity (PTI). The second branch of plant innate immunity is triggered by the recognition of specific pathogen effector proteins and known as effector-triggered immunity (ETI). Calcium (Ca(2+)) signaling is essential in both plant PTI and ETI responses. Calcium-dependent protein kinases (CDPKs) have emerged as important Ca(2+) sensor proteins in transducing differential Ca(2+) signatures, triggered by PAMPs or effectors and activating complex downstream responses. CDPKs directly transmit calcium signals by calcium binding to the elongation factor (EF)-hand domain at the C-terminus and substrate phosphorylation by the catalytic kinase domain at the N-terminus. Emerging evidence suggests that specific and overlapping CDPKs phosphorylate distinct substrates in PTI and ETI to regulate diverse plant immune responses, including production of reactive oxygen species, transcriptional reprogramming of immune genes, and the hypersensitive response.