Development of PEG-mediated genetic transformation and gene editing system of Bryum argenteum as an abiotic stress tolerance model plant.

KEY MESSAGE: To establish a sterile culture system and protoplast regeneration system for Bryum argenteum, and to establish and apply CRISPR/Cas9 system in Bryum argenteum. Bryum argenteum is a fascinating, cosmopolitan, and versatile moss species that thrives in various disturbed environments. Because of its comprehensive tolerance to the desiccation, high UV and extreme temperatures, it is emerging as a model moss for studying the molecular mechanisms underlying plant responses to abiotic stresses. However, the lack of basic tools such as gene transformation and targeted genome modification has hindered the understanding of the molecular mechanisms underlying the survival of B. argenteum in different environments. Here, we reported the protonema of B. argenteum can survive up to 95.4% water loss. In addition, the genome size of B. argenteum is approximately 313 Mb by kmer analysis, which is smaller than the previously reported 700 Mb. We also developed a simple method for protonema induction and an efficient protoplast isolation and regeneration protocol for B. argenteum. Furthermore, we established a PEG-mediated protoplast transient transfection and stable transformation system for B. argenteum. Two homologues of ABI3(ABA-INSENSITIVE 3) gene were successfully cloned from B. argenteum. To further investigate the function of the ABI3 gene in B. argenteum, we used the CRISPR/Cas9 genetic editing system to target the BaABI3A and BaABI3B gene in B. argenteum protoplasts. This resulted in mutagenesis at the target in about 2-5% of the regenerated plants. The isolated abi3a and abi3b mutants exhibited increased sensitivity to desiccation, suggesting that BaABI3A and BaABI3B play redundant roles in desiccation stress. Overall, our results provide a rapid and simple approach for molecular genetics in B. argenteum. This study contributes to a better understanding of the molecular mechanisms of plant adaptation to extreme environmental.

Drought responses and population differentiation of Calohypnum plumiforme inferred from comparative transcriptome analysis.

Bryophytes, known as poikilohydric plants, possess vegetative desiccation-tolerant (DT) ability to withstand water deficit stress. Consequently, they offer valuable genetic resources for enhancing resistance to water scarcity stress. In this research, we examined the physiological, phytohormonal, and transcriptomic changes in DT mosses Calohypnum plumiforme from two populations, with and without desiccation treatment. Comparative analysis revealed population differentiation at physiological, gene sequence, and expression levels. Under desiccation stress, the activities of superoxide dismutase (SOD) and peroxidase (POD) showed significant increases, along with elevation of soluble sugars and proteins, consistent with the transcriptome changes. Notable activation of the bypass pathway of JA biosynthesis suggested their roles in compensating for JA accumulation. Furthermore, our analysis revealed significant correlations among phytohormones and DEGs in their respective signaling pathway, indicating potential complex interplays of hormones in C plumiforme. Protein phosphatase 2C (PP2C) in the abscisic acid signaling pathway emerged as the pivotal hub in the phytohormone crosstalk regulation network. Overall, this study was one of the first comprehensive transcriptome analyses of moss C. plumiforme under slow desiccation rates, expanding our knowledge of bryophyte transcriptomes and shedding light on the gene regulatory network involved in response to desiccation, as well as the evolutionary processes of local adaptation across moss populations.

Multilevel analysis between Physcomitrium patens and Mortierellaceae endophytes explores potential long-standing interaction among land plants and fungi.

The model moss species Physcomitrium patens has long been used for studying divergence of land plants spanning from bryophytes to angiosperms. In addition to its phylogenetic relationships, the limited number of differential tissues, and comparable morphology to the earliest embryophytes provide a system to represent basic plant architecture. Based on plant-fungal interactions today, it is hypothesized these kingdoms have a long-standing relationship, predating plant terrestrialization. Mortierellaceae have origins diverging from other land fungi paralleling bryophyte divergence, are related to arbuscular mycorrhizal fungi but are free-living, observed to interact with plants, and can be found in moss microbiomes globally. Due to their parallel origins, we assess here how two Mortierellaceae species, Linnemannia elongata and Benniella erionia, interact with P. patens in coculture. We also assess how Mollicute-related or Burkholderia-related endobacterial symbionts (MRE or BRE) of these fungi impact plant response. Coculture interactions are investigated through high-throughput phenomics, microscopy, RNA-sequencing, differential expression profiling, gene ontology enrichment, and comparisons among 99 other P. patens transcriptomic studies. Here we present new high-throughput approaches for measuring P. patens growth, identify novel expression of over 800 genes that are not expressed on traditional agar media, identify subtle interactions between P. patens and Mortierellaceae, and observe changes to plant-fungal interactions dependent on whether MRE or BRE are present. Our study provides insights into how plants and fungal partners may have interacted based on their communications observed today as well as identifying L. elongata and B. erionia as modern fungal endophytes with P. patens.

Life under the snow: A year-round transcriptome analysis of Antarctic mosses in natural habitats provides insight into the molecular adaptation of plants under extreme environment.

Mosses are vital components of ecosystems, exhibiting remarkable adaptability across diverse habitats from deserts to polar ice caps. Sanionia uncinata (Hedw.) Loeske, a dominant Antarctic moss survives extreme environmental condition through perennial lifecycles involving growth and dormancy alternation. This study explores genetic controls and molecular mechanisms enabling S. uncinata to cope with seasonality of the Antarctic environment. We analysed the seasonal transcriptome dynamics of S. uncinata collected monthly from February 2015 to January 2016 in King George Island, Antarctica. Findings indicate that genes involved in plant growth were predominantly upregulated in Antarctic summer, while those associated with protein synthesis and cell cycle showed marked expression during the winter-to-summer transition. Genes implicated in cellular stress and abscisic acid signalling were highly expressed in winter. Further, validation included a comparison of the Antarctic field transcriptome data with controlled environment simulation of Antarctic summer and winter temperatures, which revealed consistent gene expression patterns in both datasets. This proposes a seasonal gene regulatory model of S. uncinate to understand moss adaptation to extreme environments. Additionally, this data set is a valuable resource for predicting genetic responses to climatic fluctuations, enhancing our knowledge of Antarctic flora's resilience to global climate change.

Plastome diversity and evolution in mosses: Insights from structural characterization, comparative genomics, and phylogenetic analysis.

Mosses play a significant role in ecology, evolution, and the economy. They belong to the nonvascular plant kingdom and are considered the closest living relatives of the first terrestrial plants. The circular chloroplast DNA molecules (plastomes) of mosses contain all the genetic information essential for chloroplast functions and represent the source of the evolutionary history of these organisms. This study comprehensively analyzed the plastomes of 47 moss species belonging to 14 orders, focusing on their size, GC content, gene loss, gene content, synteny, and evolution. The findings revealed great differences among plastome sizes, with Takakia lepidozioides (Takakiopsida) and Funaria hygrometrica (Funariales) having the largest and smallest plastomes, respectively. Moss plastomes included 69 to 89 protein-coding genes, 8 rRNA genes, and 34 to 42 tRNA genes, resulting in the total number of genes in a plastome ranging between 115 and 138. Various genes have been lost from the plastomes of different moss species, with Atrichum angustatum lacking the highest number of genes. This study also examined plastome synteny and moss evolution using comparative genomics and repeat sequence analysis. The results demonstrated that synteny and similarity levels varied across the 47 moss examined species, with some exhibiting structure similarity and others displaying structural inversions. Maximum likelihood and Bayesian approaches were used to construct a phylogenetic tree using 36 concatenated protein-coding genes, and the results revealed that the genera Sphagnum and Takakia are sister groups to the other mosses. Additionally, it was found that Tetraphidales, Polytrichales, Buxbaumiales, and Diphysciales are closely related. This research describes the evolutionary diversity of mosses and offers guidelines for future studies in this field. The findings also highlight the need for more investigations into the factors regulating plastome size variation in these plants.

Drying without dying: A genome database for desiccation-tolerant plants and evolution of desiccation tolerance.

Desiccation is typically fatal, but a small number of land plants have evolved vegetative desiccation tolerance (VDT), allowing them to dry without dying through a process called anhydrobiosis. Advances in sequencing technologies have enabled the investigation of genomes for desiccation-tolerant plants over the past decade. However, a dedicated and integrated database for these valuable genomic resources has been lacking. Our prolonged interest in VDT plant genomes motivated us to create the "Drying without Dying" database, which contains a total of 16 VDT-related plant genomes (including 10 mosses) and incorporates 10 genomes that are closely related to VDT plants. The database features bioinformatic tools, such as blast and homologous cluster search, sequence retrieval, Gene Ontology term and metabolic pathway enrichment statistics, expression profiling, co-expression network extraction, and JBrowser exploration for each genome. To demonstrate its utility, we conducted tailored PFAM family statistical analyses, and we discovered that the drought-responsive ABA transporter AWPM-19 family is significantly tandemly duplicated in all bryophytes but rarely so in tracheophytes. Transcriptomic investigations also revealed that response patterns following desiccation diverged between bryophytes and angiosperms. Combined, the analyses provided genomic and transcriptomic evidence supporting a possible divergence and lineage-specific evolution of VDT in plants. The database can be accessed at http://desiccation.novogene.com. We expect this initial release of the "Drying without Dying" plant genome database will facilitate future discovery of VDT genetic resources.

Mosses.

Often overlooked, these small but otherwise brilliant plants began covering Earth's land masses more than 450 million years ago. They saw the dinosaurs come and go, and they saw us humans coming. Mosses, liverworts and hornworts comprise the bryophytes, the second largest monophyletic clade of land plants (embryophytes), after the vascular plants (tracheophytes). Like all embryophytes, mosses exhibit a haplodiplontic life cycle. This alternation of generations (originally termed Generationswechsel in German) between the haploid gametophyte and the diploid sporophyte implies that every plant genome encodes two distinct ontogenies, in contrast to animal genomes. Contrary to tracheophytes, the haploid gametophyte is the dominant generation in mosses. Haploidy of the major tissues facilitates gene-function annotation via reverse genetics. Nevertheless, the diploid sporophyte of mosses, the spore capsule, is a visible structure unlike the gametophyte of flowering plants, which is largely reduced and embedded in the sporophyte. Visibility of both generations on one plant facilitates the analysis of the alternation of generations, and in a broader sense evo-devo studies. Whereas the conservation of moss morphology over hundreds of millions of years suggests stasis, molecular data reveal fast evolving moss genomes, leaving an enigma for evolutionary biologists. Finally, the extraordinary resilience of mosses may provide lessons for current man-made climate change. In this Primer, we will highlight some of the peculiarities of mosses from historical observations to current genomic data, with an emphasis on their development, reproduction, evolution, biotic interactions, and potential for biotechnology. Mosses from three genera - the living fossil Takakia, the ecosystems engineer Sphagnum, and the model moss Physcomitrella - exemplify the scientific insights and the applications mosses have to offer.

Comprehensive phylogenomic time tree of bryophytes reveals deep relationships and uncovers gene incongruences in the last 500 million years of diversification.

PREMISE: Bryophytes form a major component of terrestrial plant biomass, structuring ecological communities in all biomes. Our understanding of the evolutionary history of hornworts, liverworts, and mosses has been significantly reshaped by inferences from molecular data, which have highlighted extensive homoplasy in various traits and repeated bursts of diversification. However, the timing of key events in the phylogeny, patterns, and processes of diversification across bryophytes remain unclear. METHODS: Using the GoFlag probe set, we sequenced 405 exons representing 228 nuclear genes for 531 species from 52 of the 54 orders of bryophytes. We inferred the species phylogeny from gene tree analyses using concatenated and coalescence approaches, assessed gene conflict, and estimated the timing of divergences based on 29 fossil calibrations. RESULTS: The phylogeny resolves many relationships across the bryophytes, enabling us to resurrect five liverwort orders and recognize three more and propose 10 new orders of mosses. Most orders originated in the Jurassic and diversified in the Cretaceous or later. The phylogenomic data also highlight topological conflict in parts of the tree, suggesting complex processes of diversification that cannot be adequately captured in a single gene-tree topology. CONCLUSIONS: We sampled hundreds of loci across a broad phylogenetic spectrum spanning at least 450 Ma of evolution; these data resolved many of the critical nodes of the diversification of bryophytes. The data also highlight the need to explore the mechanisms underlying the phylogenetic ambiguity at specific nodes. The phylogenomic data provide an expandable framework toward reconstructing a comprehensive phylogeny of this important group of plants.

Divergent outcomes of genetic conflict on the UV sex chromosomes of Marchantia polymorpha and Ceratodon purpureus.

In species with separate sexes, the genome must produce two distinct developmental programs. Sexually dimorphic development may be controlled by either sex-limited loci or biased expression of loci transmitted through both sexes. Variation in the gene content of sex-limited chromosomes demonstrates that eukaryotic species differ markedly in the roles of these two mechanisms in governing sexual dimorphism. The bryophyte model systems Marchantia polymorpha and Ceratodon purpureus provide a particularly striking contrast. Although both species possess a haploid UV sex chromosome system, in which females carry a U chromosome and males carry a V, M. polymorpha relies on biased autosomal expression, while in C. purpureus, sex-linked genes drive dimorphism. Framing these genetic architectures as divergent outcomes of genetic conflict highlights comparative genomic analyses to better understand the evolution of sexual dimorphism.

The ancestral chromatin landscape of land plants.

Recent studies have shown that correlations between chromatin modifications and transcription vary among eukaryotes. This is the case for marked differences between the chromatin of the moss Physcomitrium patens and the liverwort Marchantia polymorpha. Mosses and liverworts diverged from hornworts, altogether forming the lineage of bryophytes that shared a common ancestor with land plants. We aimed to describe chromatin in hornworts to establish synapomorphies across bryophytes and approach a definition of the ancestral chromatin organization of land plants. We used genomic methods to define the 3D organization of chromatin and map the chromatin landscape of the model hornwort Anthoceros agrestis. We report that nearly half of the hornwort transposons were associated with facultative heterochromatin and euchromatin and formed the center of topologically associated domains delimited by protein coding genes. Transposons were scattered across autosomes, which contrasted with the dense compartments of constitutive heterochromatin surrounding the centromeres in flowering plants. Most of the features observed in hornworts are also present in liverworts or in mosses but are distinct from flowering plants. Hence, the ancestral genome of bryophytes was likely a patchwork of units of euchromatin interspersed within facultative and constitutive heterochromatin. We propose this genome organization was ancestral to land plants.

Extreme environmental adaptation mechanisms of Antarctic bryophytes are mainly the activation of antioxidants, secondary metabolites and photosynthetic pathways.

The environment in Antarctica is characterized by low temperature, intense UVB and few vegetation types. The Pohlia nutans M211 are bryophytes, which are the primary plants in Antarctica and can thrive well in the Antarctic harsh environment. The transcriptional profiling of Pohlia nutans M211 under low temperature and high UVB conditions was analyzed to explore their polar adaptation mechanism in the extreme Antarctic environment by third-generation sequencing and second-generation sequencing. In comparison to earlier second-generation sequencing techniques, a total of 43,101 non-redundant transcripts and 10,532 lncRNA transcripts were obtained, which were longer and more accurate. The analysis results of GO, KEGG, AS (alternative splicing), and WGCNA (weighted gene co-expression network analysis) of DEGs (differentially expressed genes), combined with the biochemical kits revealed that antioxidant, secondary metabolites pathways and photosynthesis were the key adaptive pathways for Pohlia nutans M211 to the Antarctic extreme environment. Furthermore, the low temperature and strong UVB are closely linked for the first time by the gene HY5 (hlongated hypocotyl 5) to form a protein interaction network through the PPI (protein-protein interaction networks) analysis method. The UVR8 module, photosynthetic module, secondary metabolites synthesis module, and temperature response module were the key components of the PPI network. In conclusion, this study will help to further explore the polar adaptation mechanism of Antarctic plants represented by bryophytes and to enrich the polar gene resources.

Molecular characterization of a moss isoprene synthase provides insight into its evolution.

This is the first report on the molecular characterization of isoprene synthase (ISPS) from the moss Calohypnum plumiforme. After isoprene emission from C. plumiforme was confirmed, the cDNA encoding C. plumiforme ISPS (CpISPS) was narrowed down using a genome database associated with protein structure prediction, and a CpISPS gene was identified. The recombinant CpISPS, produced in Escherichia coli, converted dimethylallyl diphosphate to isoprene. Phylogenetic analysis indicated similarity between the amino acid sequences of CpISPS and moss diterpene cyclases (DTCs) but not ISPSs of higher plants, implying that CpISPS is derived from moss DTCs and is evolutionarily unrelated to canonical ISPSs of higher plants. CpISPS is a novel class I cyclase of the terpene synthase-c subfamily harboring αß domains. This study will help further study of isoprene biosynthesis and the physiological functions of isoprene in mosses.

The putative myristoylome of Physcomitrium patens reveals conserved features of myristoylation in basal land plants.

KEYMESSAGE: The putative myristoylome of moss P. patens opens an avenue for studying myristoylation substrates in non-canonical model plants. A myristoylation signal was shown sufficient for membrane targeting and useful for membrane dynamics visualization during cell growth. N-myristoylation (MYR) is one form of lipid modification catalyzed by N-myristoyltransferase that enables protein-membrane association. MYR is highly conserved in all eukaryotes. However, the study of MYR is limited to a few models such as yeasts, humans, and Arabidopsis. Here, using prediction tools, we report the characterization of the putative myristoylome of the moss Physcomitrium patens. We show that basal land plants display a similar signature of MYR to Arabidopsis and may have organism-specific substrates. Phylogenetically, MYR signals have mostly co-evolved with protein function but also exhibit variability in an organism-specific manner. We also demonstrate that the MYR motif of a moss brassinosteroid-signaling kinase is an efficient plasma membrane targeting signal and labels lipid-rich domains in tip-growing cells. Our results provide insights into the myristoylome in a basal land plant and lay the foundation for future studies on MYR and its roles in plant evolution.

Genome-Wide Analysis of Long Non-Coding RNAs Related to UV-B Radiation in the Antarctic Moss Pohlia nutans.

Antarctic organisms are consistently suffering from multiple environmental pressures, especially the strong UV radiation caused by the loss of the ozone layer. The mosses and lichens dominate the vegetation of the Antarctic continent, which grow and propagate in these harsh environments. However, the molecular mechanisms and related regulatory networks of these Antarctic plants against UV-B radiation are largely unknown. Here, we used an integrated multi-omics approach to study the regulatory mechanism of long non-coding RNAs (lncRNAs) of an Antarctic moss (Pohlia nutans) in response to UV-B radiation. We identified a total of 5729 lncRNA sequences by transcriptome sequencing, including 1459 differentially expressed lncRNAs (DELs). Through functional annotation, we found that the target genes of DELs were significantly enriched in plant-pathogen interaction and the flavonoid synthesis pathway. In addition, a total of 451 metabolites were detected by metabonomic analysis, and 97 differentially change metabolites (DCMs) were found. Flavonoids account for 20% of the total significantly up-regulated metabolites. In addition, the comprehensive transcriptome and metabolome analyses revealed the co-expression pattern of DELs and DCMs of flavonoids. Our results provide insights into the regulatory network of lncRNA under UV-B radiation and the adaptation of Antarctic moss to the polar environments.

Transcriptome Reveals the Molecular Mechanism of the ScALDH21 Gene from the Desert Moss Syntrichia caninervis Conferring Resistance to Salt Stress in Cotton.

The desert moss Syntrichia caninervis has proven to be an excellent plant material for mining resistance genes. The aldehyde dehydrogenase 21 (ScALDH21) gene from S. caninervis has been shown to confer tolerance to salt and drought, but it is unclear how the transgene ScALDH21 regulates tolerance to abiotic stresses in cotton. In the present work, we studied the physiological and transcriptome analyses of non-transgenic (NT) and transgenic ScALDH21 cotton (L96) at 0 day, 2 days, and 5 days after salt stress. Through intergroup comparisons and a weighted correlation network analysis (WGCNA), we found that there were significant differences between NT and L96 cotton in the plant hormone, Ca2+, and mitogen-activated protein kinase (MAPK) signaling pathways as well as for photosynthesis and carbohydrate metabolism. Overexpression of ScALDH21 significantly increased the expression of stress-related genes in L96 compared to NT cotton under both normal growth and salt stress conditions. These data suggest that the ScALDH21 transgene can scavenge more reactive oxygen species (ROS) in vivo relative to NT cotton and improve cotton resistance to salt stress by increasing the expression of stress-responsive genes, responding quickly to stress stimuli, enhancing photosynthesis and improving carbohydrate metabolism. Therefore, ScALDH21 is a promising candidate gene to improve resistance to salt stress, and the application of this gene in cotton provides new insights into molecular plant breeding.

An optimized transformation protocol for Anthoceros agrestis and three more hornwort species.

Land plants comprise two large monophyletic lineages, the vascular plants and the bryophytes, which diverged from their most recent common ancestor approximately 480 million years ago. Of the three lineages of bryophytes, only the mosses and the liverworts are systematically investigated, while the hornworts are understudied. Despite their importance for understanding fundamental questions of land plant evolution, they only recently became amenable to experimental investigation, with Anthoceros agrestis being developed as a hornwort model system. Availability of a high-quality genome assembly and a recently developed genetic transformation technique makes A. agrestis an attractive model species for hornworts. Here we describe an updated and optimized transformation protocol for A. agrestis, which can be successfully used to genetically modify one more strain of A. agrestis and three more hornwort species, Anthoceros punctatus, Leiosporoceros dussii, and Phaeoceros carolinianus. The new transformation method is less laborious, faster, and results in the generation of greatly increased numbers of transformants compared with the previous method. We have also developed a new selection marker for transformation. Finally, we report the development of a set of different cellular localization signal peptides for hornworts providing new tools to better understand the hornwort cell biology.

Proteomic and Transcriptomic Responses of the Desiccation-Tolerant Moss Racomitrium canescens in the Rapid Rehydration Processes.

The moss Racomitrium canescens (R. canescens) has strong desiccation tolerance. It can remain desiccated for years and yet recover within minutes of rehydration. Understanding the responses and mechanisms underlying this rapid rehydration capacity in bryophytes could identify candidate genes that improve crop drought tolerance. We explored these responses using physiology, proteomics, and transcriptomics. Label-free quantitative proteomics comparing desiccated plants and samples rehydrated for 1 min or 6 h suggesting that damage to chromatin and the cytoskeleton had occurred during desiccation, and pointing to the large-scale degradation of proteins, the production of mannose and xylose, and the degradation of trehalose immediately after rehydration. The assembly and quantification of transcriptomes from R. canescens across different stages of rehydration established that desiccation was physiologically stressful for the plants; however, the plants recovered rapidly once rehydrated. According to the transcriptomics data, vacuoles appear to play a crucial role in the early stages of R. canescens recovery. Mitochondria and cell reproduction might recover before photosynthesis; most biological functions potentially restarted after ~6 h. Furthermore, we identified novel genes and proteins related to desiccation tolerance in bryophytes. Overall, this study provides new strategies for analyzing desiccation-tolerant bryophytes and identifying candidate genes for improving plant drought tolerance.

The phylogeny of Syntrichia: An ecologically diverse clade of mosses with an origin in South America.

PREMISE: To address the biodiversity crisis, we need to understand the evolution of all organisms and how they fill geographic and ecological space. Syntrichia is one of the most diverse and dominant genera of mosses, ranging from alpine habitats to desert biocrusts, yet its evolutionary history remains unclear. METHODS: We present a comprehensive phylogenetic analysis of Syntrichia, based on both molecular and morphological data, with most of the named species and closest outgroups represented. In addition, we provide ancestral-state reconstructions of water-related traits and a global biogeographic analysis. RESULTS: We found 10 major well-resolved subclades of Syntrichia that possess geographical or morphological coherence, in some cases representing previously accepted genera. We infer that the extant species diversity of Syntrichia likely originated in South America in the early Eocene (56.5-43.8 million years ago [Mya]), subsequently expanded its distribution to the neotropics, and finally dispersed to the northern hemisphere. There, the clade experienced a recent diversification (15-12 Mya) into a broad set of ecological niches (e.g., the S. caninervis and S. ruralis complexes). The transition from terricolous to either saxicolous or epiphytic habitats occurred more than once and was associated with changes in water-related traits. CONCLUSIONS: Our study provides a framework for understanding the evolutionary history of Syntrichia through the combination of morphological and molecular characters, revealing that migration events that shaped the current distribution of the clade have implications for morphological character evolution in relation to niche diversity.

Functional identification of moss PpGATA1 provides insights into the evolution of LLM-domain B-GATA transcription factors in plants.

B-GATA transcription factors with the LLM domain (LLM-domain B-GATAs) play important roles in developmental processes and environmental responses in flowering plants. Their characterization can therefore provide insights into the structural and functional evolution of functional gene families. Phylogenetic and sequence analysis suggests that LLM-domain B-GATAs evolved from ancestral GATA transcription factors before the divergence of chlorophyte algae and Streptophyta. We compared the function of PpGATA1, a LLM-domain B-GATA gene in moss Physcomitrium patens, with Arabidopsis thaliana counterparts and showed that, in P. patens, PpGATA1 controls growth and greening in haploid gametophytes, while in transgenic Arabidopsis it affects germination, leaf development, flowering time, greening and light responses in diploid sporophytes. These PpGATA1 functions are similar to those of Arabidopsis counterparts, AtGNC, AtGNL and AtGATA17. PpGATA1 was able to complement the role of GNC and GNL in a gnc gnl double mutant, and the LLM domains of PpGATA1 and GNC behaved similarly. The functions of LLM-domain B-GATAs regulating hypocotyl elongation and cotyledon epinasty in flowering plants pre-exist before the divergence of mosses and the lineage leading to flowering plants. This study sheds light on adaption of PpGATA1 and its homologs to new developmental designs during the evolution.

A comparison of drying methods on the quality for bryophyte molecular specimens collected in the field.

A major challenge in extracting high-quality DNA from bryophytes is the treatment of bryophyte material in the field. The existing and commonly used treatment methods in the field have several shortcomings. Natural drying methods can lead to DNA breaks. In addition, it is highly cumbersome to carry large quantities of silica gel in the field due to its weight and high risk of contamination among samples. In this study, we explored more convenient drying methods to treat bryophyte specimens and promote more efficient DNA recovery. The quantity and quality of genomic DNA extracted from every bryophyte species using different drying methods, including hot-air drying methods (150°C, 80°C, and 40°C), natural drying method, and silica gel drying method, were measured. Spectrophotometry, electrophoresis, and PCR amplification were performed to assess the effects of different drying methods. The results of total DNA purity, total DNA concentration, PCR success, and OD 260/230 ratios suggested that the hot-air drying (40-80°C) was the best method. The morphological comparison revealed that hot-air drying at 40°C and 80°C exerted no significant adverse effects on plant morphology and taxonomic studies. Thus, this method prevents rapid DNA degradation and silica gel pollution and saves the workforce from carrying large amounts of silica gel to the field. Several inexpensive devices, such as portable hairdryers, fan heaters, and electric blankets, are available that can be easily carried to the field for drying molecular specimens.