Automated, High-Throughput Protoplast Transfection for Gene Editing and Transgene Expression Studies.

In an era of cost-efficient gene synthesis and high-throughput construct assembly, the onus of scientific experimentation is on the rate of in vivo testing for the identification of top performing candidates or designs. Assay platforms that are relevant to the species of interest and in the tissue of choice are highly desirable. A protoplast isolation and transfection method that is compatible with a large repertoire of species and tissues would be the platform of choice.

CRISPR/LbCas12a-Mediated Genome Editing in Soybean.

Currently methods for generating soybean edited lines are time-consuming, inefficient, and limited to certain genotypes. Here we describe a fast and highly efficient genome editing method based on CRISPR-Cas12a nuclease system in soybean. The method uses Agrobacterium-mediated transformation to deliver editing constructs and uses aadA or ALS genes as selectable marker.

Cellular Stress Upregulates Indole Signaling Metabolites in Escherichia coli.

Escherichia coli broadly colonize the intestinal tract of humans and produce a variety of small molecule signals. However, many of these small molecules remain unknown. Here, we describe a family of widely distributed bacterial metabolites termed the "indolokines.

Hierarchical and Dynamic Regulation of Defense-Responsive Specialized Metabolism by WRKY and MYB Transcription Factors.

The plant kingdom produces hundreds of thousands of specialized bioactive metabolites, some with pharmaceutical and biotechnological importance. Their biosynthesis and function have been studied for decades, but comparatively less is known about how transcription factors with overlapping functions and contrasting regulatory activities coordinately control the dynamics and output of plant specialized metabolism. Here, we performed temporal studies on pathogen-infected intact host plants with perturbed transcription factors.

Expansion of a core regulon by transposable elements promotes Arabidopsis chemical diversity and pathogen defense.

Plants synthesize numerous ecologically specialized, lineage-specific metabolites through biosynthetic gene duplication and functional specialization. However, it remains unclear how duplicated genes are wired into existing regulatory networks. We show that the duplicated gene CYP82C2 has been recruited into the WRKY33 regulon and indole-3-carbonylnitrile (ICN) biosynthetic pathway through exaptation of a retroduplicated LINE retrotransposon (EPCOT3) into an enhancer.

Evolution of Glucosinolate Diversity via Whole-Genome Duplications, Gene Rearrangements, and Substrate Promiscuity.

Over several decades, glucosinolates have become a model system for the study of specialized metabolic diversity in plants. The near-complete identification of biosynthetic enzymes, regulators, and transporters has provided support for the role of gene duplication and subsequent changes in gene expression, protein function, and substrate specificity as the evolutionary bases of glucosinolate diversity. Here, we provide examples of how whole-genome duplications, gene rearrangements, and substrate promiscuity potentiated the evolution of glucosinolate biosynthetic enzymes, regulators, and transporters by natural selection.

A new cyanogenic metabolite in Arabidopsis required for inducible pathogen defence.

Thousands of putative biosynthetic genes in Arabidopsis thaliana have no known function, which suggests that there are numerous molecules contributing to plant fitness that have not yet been discovered. Prime among these uncharacterized genes are cytochromes P450 upregulated in response to pathogens. Here we start with a single pathogen-induced P450 (ref.

Seed after-ripening and dormancy determine adult life history independently of germination timing.

• Seed dormancy can affect life history through its effects on germination time. Here, we investigate its influence on life history beyond the timing of germination. • We used the response of Arabidopsis thaliana to chilling at the germination and flowering stages to test the following: how seed dormancy affects germination responses to the environment; whether variation in dormancy affects adult phenology independently of germination time; and whether environmental cues experienced by dormant seeds have an effect on adult life history.