The regulation of stomatal lineage cell development has been extensively investigated. However a comprehensive characterization of this biological process based on single-cell transcriptome analysis has not yet been reported. Here, we performed RNA-seq on over 12,844 individual cells from the cotyledons of five-day-old Arabidopsis seedlings. We identified 11 cell clusters corresponding mostly to cells at specific stomatal developmental stages with a series of new marker genes. Comparative analysis of genes with the highest variable expression in these cell clusters revealed three transcriptional networks that regulate the development of mesophyll and guard cells, as well as the differentiation from protodermal to guard mother cells. We investigated the developmental dynamics of marker genes via pseudo-time analysis which revealed potential interactions between them. The identification of several novel marker genes suggests new regulatory mechanisms during development of stomatal cell lineage. .

Recent advances in carboxylic acid reductases and their practical applications in bio-cascade processes.

Carotenoids are important pigments in plants that play crucial roles in plant growth and in plant responses to environmental stress. Lycopene β cyclase (β-LCY) functions at the branch point of the carotenoid biosynthesis pathway, catalyzing the cyclization of lycopene. Here, a β-LCY gene from Nicotiana tabacum, designated as Ntβ-LCY1, was cloned and functionally characterized. Robust expression of Ntβ-LCY1 was found in leaves, and Ntβ-LCY1 expression was obviously induced by salt, drought, and exogenous abscisic acid treatments. Strong accumulation of carotenoids and expression of carotenoid biosynthesis genes resulted from Ntβ-LCY1 overexpression. Additionally, compared to wild-type plants, transgenic plants with overexpression showed enhanced tolerance to salt and drought stress with higher abscisic acid levels and lower levels of malondialdehyde and reactive oxygen species. Conversely, transgenic RNA interference plants had a clear albino phenotype in leaves, and some plants did not survive beyond the early developmental stages. The suppression of Ntβ-LCY1 expression led to lower expression levels of genes in the carotenoid biosynthesis pathway and to reduced accumulation of carotenoids, chlorophyll, and abscisic acid. These results indicate that Ntβ-LCY1 is not only a likely cyclization enzyme involved in carotenoid accumulation but also confers salt and drought stress tolerance in Nicotiana tabacum.

BackgroundThe CRISPR/Cas9 system is being used for genome editing purposes by many research groups in multiple plant species. Traditional sequencing methods to identify homozygous mutants are time-consuming, laborious and expensive.ResultsWe have developed a method to screen CRISPR/Cas9-induced mutants through Mutation Sites Based Specific Primers Polymerase Chain Reaction (MSBSP-PCR). The MSBSP-PCR method was successfully used to identify homozygous/biallelic mutants in Nicotiana tabacum and Arabidopsis thaliana, and we speculate that it can be used for the identification of CRISPR/Cas9-induced mutants in other plant species. Compared to traditional sequencing methods, MSBSP-PCR is simpler, faster and cheaper.ConclusionsThe MSBSP-PCR method is simple to implement and can save time and cost in the screening of CRISPR/Cas9-induced homozygous/biallelic mutants.Electronic supplementary materialThe online version of this article (10.1186/s13007-018-0305-8) contains supplementary material, which is available to authorized users.

No abstract

Veratramine (VAM) is the major lipid-soluble alkaloid existing in Veratrum nigrum L. that has been demonstrated to exert neurotoxic effects. To better understand the potential mechanism of neurotoxicity of VAM, VAM-induced DNA damage was measured in the cerebellum and cerebral cortex of mice after a 7-day repetitive oral dose by using single-cell gel electrophoresis (comet assay). A method based on high-performance liquid chromatography-electrospray ionization tandem mass spectrometry was developed for the determination of VAM and its in vivo and in vitro metabolites, to establish the potential correlation between metabolites and neurotoxicity. In vitro experiment was carried out using rat liver microsomes, whereas the in vivo study was conducted on rats at a single dose of 3 mg/kg. The results showed that VAM caused DNA damage in the cerebellum and cerebral cortex of mice in a dose-dependent manner. Phenyl mono-oxidation, sulfate conjugation and phenyl di-oxidation were proposed to be the main in vivo metabolic pathways of VAM, whereas the major in vitro metabolic pathways were phenyl mono-oxidation, hydroxylation and methylation. Phenyl-oxidation reaction was likely to be associated with reactive oxygen species production, leading to the DNA damage in the mouse brain.

Previous investigations have shown that the SUPPRESSORS OF MAX2 1-LIKE6, 7 and 8 (SMXL6, 7 and 8) proteins redundantly repress strigolactone (SL) signaling in plant growth and development. Recently, a growing body of evidence indicated that SLs positively regulate plant drought resistance through functional analyses of genes involved in SL biosynthesis and positive regulation of SL signaling. However, the functions of the SL-signaling negative regulators SMXL6, 7 and 8 in drought resistance and the associated mechanisms remain elusive. To reveal the functions of these SMXL proteins, we analyzed the drought-resistant phenotype of the triple smxl6,7,8 mutant plants and studied several drought resistance-related traits. Our results showed that the smxl6,7,8 mutant plants were more resistant to drought than wild-type plants. Physiological investigations indicated that the smxl6,7,8 mutant plants exhibited higher leaf surface temperature, reduced cuticle permeability, as well as decreases in drought-induced water loss and cell membrane damage in comparison with wild-type plants. Additionally, smxl6,7,8 mutant plants displayed an increase in anthocyanin biosynthesis during drought, enhanced detoxification capacity and increased sensitivity to abscisic acid in cotyledon opening and growth inhibition assays. A good correlation between the expression levels of some relevant genes and the examined physiological and biochemical traits was observed. Our findings together indicate that the SMXL6, 7 and 8 act as negative regulators of drought resistance, and that disruption of these SMXL genes in crops may provide a novel way to improve their drought resistance.

Chiral amino alcohols are essential building blocks for the pharmaceutical industry, and are widely presented in natural and synthetic bioactive compounds. Amine dehydrogenases (AmDHs) can asymmetrically reduce prochiral ketones with...