Molecule fluorescent probes for sensing and imaging analytes in plants: Developments and challenges

The subgenus Cerasus, one of the most important groups in the genus Prunus sensu lato, comprises over 100 species; however, the taxonomic classification and phylogenetic relationships of Cerasus remain controversial. Therefore, it is necessary to reconstruct the phylogenetic tree for known Cerasus species. Here, we report the chloroplast (cp) genome sequences of 11 Cerasus species to provide insight into evolution of the plastome. The cp genomes of the 11 Cerasus species (157,571–158,830 bp) displayed a typical quadripartite circular structure. The plastomes contain 115 unique genes, including 80 protein-coding genes, four ribosomal RNAs, and 31 transfer RNAs. Twenty genes were found to be duplicated in inverted repeats as well as at the boundary. The conserved non-coding sequences showed significant divergence compared with the coding regions. We found 12 genes and 14 intergenic regions with higher nucleotide diversity and more polymorphic sites, including matK, rps16, rbcL, rps16-trnQ, petN-psbM, and trnL-trnF. During cp plastome evolution, the codon profile has been strongly biased toward the use of A/T at the third base, and leucine and isoleucine codons appear the most frequently. We identified strong purifying selection on the rpoA, cemA, atpA, and petB genes; whereas ccsA, rps19, matK, rpoC2, ycf2 and ndhI showed a signature of possible positive selection during the course of Cerasus evolution. In addition, we further analyzed the phylogenetic relationships of these species with 57 other congenic related species.Through reconstructing the Cerasus phylogeny tree, we found that true cherry is similar to the flora of China forming a distinct group, from which P. mahaleb was separated as an independent subclade. Microcerasus was genetically closer to Amygdalus, Armeniaca, and Prunus (sensu stricto) than to members of true cherry, whereas P. japonica and P. tomentosa were most closely related to P. triloba and P. pedunculata. However, P. tianshanica formed a clade with P. cerasus, P. fruticosa, P. cerasus × P. canescens ‘Gisela 6’, and P. avium as a true cherry group. These results provide new insights into the plastome evolution of Cerasus, along with potential molecular markers and candidate DNA barcodes for further phylogenetic and phylogeographic analyses of Cerasus species.

https://www.frontiersin.org/articles/10.3389/fpls.2023.1070600/pdf

Evolutionary and phylogenetic analyses of 11 Cerasus species based on the complete chloroplast genome

Osmotic stress caused by low-temperature, drought and salinity was a prevalent abiotic stress in plant that severely inhibited plant development and agricultural yield, particularly in tea plant. Jasmonic acid (JA) is an important phytohormone involving in plant stress. However, underlying molecular mechanisms of JA modulated osmotic stress response remains unclear. In this study, high concentration of mannitol induced JA accumulation and increase of peroxidase activity in tea plant. Integrated transcriptome mined a JA signaling master, MYC2 transcription factor is shown as a hub regulator that induced by mannitol, expression of which positively correlated with JA biosynthetic genes (LOX and AOS) and peroxidase genes (PER). CsMYC2 was determined as a nuclei-localized transcription activator, furthermore, Protein-DNA interaction analysis indicated that CsMYC2 was positive regulator that activated the transcription of CsLOX7, CsAOS2, CsPER1 and CsPER3 via bound with their promoters, respectively. Suppression of CsMYC2 expression resulted in a reduced JA content and peroxidase activity and osmotic stress tolerance of tea plant. Overexpression of CsMYC2 in Arabidopsis improved JA content, peroxidase activity and plants tolerance against mannitol stress. Together, we proposed a positive feedback loop mediated by CsMYC2, CsLOX7 and CsAOS2 which constituted to increase the tolerance of osmotic stress through fine-tuning the accumulation of JA levels and increase of POD activity in tea plant. 

JA-mediated MYC2/LOX/AOS feedback loop regulates osmotic stress response in tea plant (Camellia sinensis)

Among the New Plant Breeding Techniques (NPBTs), the CRISPR/Cas9 system represents a useful tool for target gene editing, improving the traits of the plants rapidly. This technology allows targeting one or more sequences simultaneously, as well as introducing new genetic variations by homology-directed recombination. However, the technology of CRISPR/Cas9 remains a challenge for some polyploid woody species, since all the different alleles for which the mutation is required must be simultaneously targeted. In this work we describe improved protocols adapting the CRISPR/Cas9 system to highbush blueberry (Vaccinium corymbosum L.), using Agrobacterium-mediated transformation. As a proof of concept, we targeted the gene encoding for phytoene desaturase, whose mutation disrupts chlorophyll biosynthesis allowing for the visual assessment of knockout efficiency. Leaf explants of in vitro-cultured blueberry cv. Berkeley has been transformed with a CRISPR/Cas9 construct containing two guide RNAs (gRNA1 and gRNA2) targeting two conserved gene regions of pds and subsequently maintained on a selection medium enriched with kanamycin. After 4 weeks in culture on the selection medium, the kanamycin-resistant lines were isolated, and the genotyping of these lines through Sanger sequencing revealed successful gene editing. Some of mutant shoot lines included albino phenotypes, even if the editing efficiencies were quite low for both gRNAs, ranging between 2.1 and 9.6% for gRNA1 and 3.0 and 23.8 for gRNA2. Here we showed a very effective adventitious shoot regeneration protocol for the commercial cultivar of highbush blueberry “Berkeley”, and a further improvement in the use of CRISPR/Cas9 system in Vaccinium corymbosum L., opening the way to the breeding mediated by biotechnological approaches.

/pdf/knockout-of-phytoene-desaturase-gene-using-crispr-cas9-in-hxe6uovj.pdf

Knockout of phytoene desaturase gene using CRISPR/Cas9 in highbush blueberry

Melatonin (N-acetyl-5-hydroxy tryptamine), a multipotent biomolecule is well known for its ability to confer tolerance to several abiotic and biotic stresses. The regulation of melatonin-mediated drought tolerance in drought-distinguished varieties can be different due to discriminating redox levels. The present study was focused on assessing the effects of melatonin priming against polyethylene glycol (PEG)-induced stress with respect to the antioxidant system, photosynthetic parameters, lipoxygenase expression, JA and ABA levels in drought-sensitive (Kadiri-7) and drought-tolerant (Kadiri-9) varieties. Exogenous melatonin alleviated the drought stress effects in sensitive variety (Kadiri-7) by increasing the endogenous melatonin content with an improved antioxidant system and photosynthetic attributes. The primed stressed plants of the sensitive variety exhibited reduced expression and activity of the chlorophyll degrading enzymes (Chl-deg PRX, pheophytinase and chlorophyllase) with a concomitant increase in chlorophyll content in comparison to unprimed controls. Interestingly, melatonin priming stimulated higher expression and activity of lipoxygenase (LOX) as well as enhanced the expression of genes involved in the synthesis of jasmonic acid (JA) including its content in drought stressed plants of the sensitive variety. The expression of NCED3 (involved in ABA-biosynthesis) was upregulated while CYP707A2 (ABA-degradation) was downregulated which corresponded with higher ABA levels. Contrastingly, priming caused a decrease in endogenous melatonin content under drought stress in tolerant variety (Kadiri-9) which might be due to feedback inhibition of its synthesis to maintain intracellular redox balance and regulate better plant metabolism. Furthermore, the higher endogenous melatonin content along with improved antioxidant system, photosynthetic efficiency and LOX expression associated with the increased levels of JA and ABA in unprimed stressed plants of the tolerant variety (Kadiri-9) is pointing towards the effectiveness of melatonin in mediating drought stress tolerance. Overall, exogenous melatonin alleviated the adverse effects of drought stress in sensitive variety while having no add-on effect on drought stress responses of tolerant variety which is inherently equipped to withstand the given duration of drought stress treatment.

Melatonin induces drought tolerance by modulating lipoxygenase expression, redox homeostasis and photosynthetic efficiency in Arachis hypogaea L

Effects of cellulose nanofibrils treatment on antioxidant properties and aroma of fresh-cut apples.

Hydrogen peroxide (H2O2), as a main component of reactive oxygen species (ROS), serves as a key signaling molecule relevant to plant stress response and health status. Many strategies have been developed for detecting or quantifying H2O2 concentration. However, reports on simply, visibly tracking H2O2 fluctuation in vivo are limited. Here, for visibly tracking the plant H2O2 wave, a green fluorescent phenotypic probe was designed by merging a H2O2-sensitive tertiary amine moiety with the core fluorophore tetraphenylethene skeleton. The green fluorescence emission is quenched up to 52% by H2O2 with good sensitivity, selectivity, and reversibility within the plant physiological range of 10-100 μM H2O2. In response to various abiotic stresses, including mechanical damage, high salt, strong light and drought, fluorescence fluctuations, response to H2O2 concentration alterations in vivo was visible to the naked eye under irradiation of commercially available UV light (365 nm) after simple injection of this H2O2 probe solution into seedling leaves. This phenotypic fluorescent H2O2 probe illustrates great potential as early sensors of plant health under stress without the aid of skillful operation and specialized equipment.

Simple Phenotypic Sensor for Visibly Tracking H2O2 Fluctuation to Detect Plant Health Status.

Abstract Apple (Malus\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{upgreek} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} $ \times $\end{document}domestica) is a popular temperate fruit crop worldwide. However, its growth, productivity, and quality are often adversely affected by abiotic stresses such as drought, extreme temperature, and high salinity. Due to the long juvenile phase and highly heterozygous genome, the conventional breeding approaches for stress-tolerant cultivars are time-consuming and resource-intensive. These issues may be resolved by feasible molecular breeding techniques for apples, such as gene editing and marker-assisted selection. Therefore, it is necessary to acquire a more comprehensive comprehension of the molecular mechanisms underpinning apples’ response to abiotic stress. In this review, we summarize the latest research progress in the molecular response of apples to abiotic stressors, including the gene expression regulation, protein modifications, and epigenetic modifications. We also provide updates on new approaches for improving apple abiotic stress tolerance, while discussing current challenges and future perspectives for apple molecular breeding.

Insights into the molecular mechanisms underlying responses of apple trees to abiotic stresses

Functional identification of apple Baby Boom in genetic transformation and somatic embryogenesis

Genome-wide identification of apple auxin receptor family genes and functional characterization of MdAFB1

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