Biology subjects

Michael, T. P.

Publications and source records attributed to Michael, T. P..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Chromosome-level baobab (Adansonia digitata) genome illuminates its evolutionary insights

AbstractBaobab, Adansonia digitata, is a long-lived tree endemic to Africa that holds great economic, ecological, and cultural value. However, our knowledge of its genomic features, evolutionary history, and diversity is limited, rendering it orphaned scientifically. We generated a haploid chromosome-level reference genome anchored into 42 chromosomes for A. digitata, as well as draft assemblies for a sibling tree, two trees from distinct locations in Africa, and a related species, A. za from Madagascar. Unlike any other plant to date, DNA transposable elements (TEs) make up 33% of the A. digitata genome compared to only 10% long terminal repeat retrotransposons (LTR-RTs), which are usually predominant in plant genomes. Baobab has undergone a whole genome duplication (WGD) shared with the Malvoideae [~]30 million years ago (MYA), as well as a confirmed autotetraplody event 3-4 million MYA that coincides with the most recent burst of TE insertions. Resequencing 25 A. digitata trees from Africa revealed three subpopulations that suggest gene flow through most of West Africa but separated from East Africa. Gene enrichment analysis for baobab-specific and high fixation index (Fst) suggested baobab may have retained multiple copies of circadian, light and growth genes to coordinate genome protection for longevity through the UV RESISTANCE LOCUS 8 (UVR8) and synchronizing flower development with pollinators. This study lays the groundwork for the creation of breeding resources and the conservation of baobab biodiversity.

genomics↗

bioRxiv2024

Telomere Length in Plants Estimated with Long Read Sequencing

Telomeres play an important role in chromosome stability and their length is thought to be related to an organisms lifestyle and lifespan. Telomere length is variable across plant species and between cultivars of the same species, possibly conferring adaptive advantage. However, it is not known whether telomere length is related to lifestyle or life span across a diverse array of plant species due to the lack of information on telomere length in plants. Here we leverage genomes assembled with long read sequencing data to estimate telomere length by chromosome. We find that long read assemblies based on Oxford Nanopore Technologies (ONT) accurately predict telomere length in the two model plant species Arabidopsis thaliana and Oryza sativa matching lab-based length estimates. We then estimate telomere length across an array of plant species with different lifestyles and lifespans and find that in general gymnosperms have shorter telomeres compared to eudicots and monocots. Crop species frequently have longer telomeres than their wild relatives, and species that have been maintained clonally such as hemp have long telomeres possibly reflecting that this lifestyle requires long term chromosomal stability.

plant biology↗

bioRxiv2024

Genome-Wide Association Study Reveals Influence of Cell-specific Gene Networks on Soybean Root System Architecture

Root system architecture (RSA) describes the shape and arrangement of a plants roots in the soil including the angle, rate of growth, and type of individual roots, which facilitates the uptake of nutrients and water. In crop improvement efforts, RSA has been less well studied due to the technical challenges associated with phenotyping roots as well as a focus on above-ground traits such as yield. We developed a gel-based root phenotyping system called RADICYL (Root Architecture 3D Cylinder), which is a non-invasive, high-throughput approach that enabled us to measure 15 RSA traits. We leveraged RADICYL to perform a comprehensive genome-wide association study (GWAS) with a panel of 371 diverse soybean elite lines, cultivars, landraces, and closely related species to identify gene networks underlying RSA. We identified 54 significant single nucleotide polymorphisms (SNPs) in our GWAS, some of which were shared across multiple RSA traits while others were specific to a given trait. We generated a single cell atlas of the soybean root using single nuclei RNA sequencing (snRNAseq) to explore the associated genes in the context of root tissues. Using gene co-expression network (GCN) analyses applied to RNA-seq of soybean root tissues, we identified network-level associations of genes predominantly expressed in endodermis with root width, and of those expressed in metaphloem with lateral root length. Our results suggest that pathways active in the endodermis and metaphloem cell-types influence soybean root system architecture.

plant biology↗

bioRxiv2023

The genomes and epigenomes of aquatic plants (Lemnaceae) promote triploid hybridization and clonal reproduction

The Lemnaceae (duckweeds) are the worlds smallest but fastest growing flowering plants. Prolific clonal propagation facilitates continuous micro-cropping for plant-based protein and starch production, and holds tremendous promise for sequestration of atmospheric CO2. Here, we present chromosomal assemblies, annotations, and phylogenomic analysis of Lemna genomes that uncover candidate genes responsible for the metabolic and developmental traits of the family, such as anatomical reduction, adaxial stomata, lack of stomatal closure, and carbon sequestration via crystalline calcium oxalate. Lemnaceae have selectively lost genes required for RNA interference, including Argonaute genes required for reproductive isolation (the triploid block) and haploid gamete formation. Triploid hybrids arise commonly among Lemna, and we have found mutations in highly-conserved meiotic crossover genes that could support polyploid meiosis. Syntenic comparisons with Wolffia and Spirodela reveal that diversification of these genera coincided with the "Azolla event" in the mid-Eocene, during which aquatic macrophytes reduced high atmospheric CO2 levels to those of the current ice age. Facile regeneration of transgenic fronds from tissue culture, aided by reduced epigenetic silencing, makes Lemna a powerful biotechnological platform, as exemplified by recent engineering of high-oil Lemna that outperforms oil seed crops.

plant biology↗

bioRxiv2023

Subgenome dominance shapes novel gene evolution in the decaploid pitcher plant Nepenthes gracilis

Subgenome dominance after whole-genome duplication generates distinction in gene number and expression at the level of chromosome sets, but it remains unclear how this process may be involved in evolutionary novelty. Here, we generated a chromosome-scale genome assembly of the Asian pitcher plant Nepenthes gracilis to analyze how its novel traits (dioecy and carnivorous pitcher leaves) are linked to genomic evolution. We found a decaploid karyotype with five complete sets of syntenic chromosomes (2n = 10x = 80) yet with a clear indication of subgenome dominance and highly diploidized gene contents. The male-linked and pericentromerically located region on the putative sex chromosome was identified in a recessive subgenome and was found to harbor three transcription factors involved in flower and pollen development, including a likely neofunctionalized LEAFY duplicate. Transcriptomic and syntenic analyses of carnivory-related genes suggested that the paleopolyploidization events seeded genes that subsequently formed tandem clusters in recessive subgenomes with specific expression in the digestive zone of the pitcher, where specialized cells digest prey and absorb derived nutrients. Novel gene evolution in recessive subgenomes is likely to be prevalent because duplicates were enriched with Nepenthes-specific genes with tissue-specific expression, including those expressed in trapping pitchers. Thus, subgenome dominance likely contributed to evolutionary novelty by allowing recessive subgenomes experiencing relaxed purifying selection to serve as a preferred host of novel tissue-specific duplicates. Our results provide insight into how polyploids, which may frequently be evolutionary dead-ends, have given rise to novel traits in exceptionally thriving high-ploidy lineages.

plant biology↗

bioRxiv2023

Genetic Mapping of SNP Markers and Candidate Genes Associated with Day-Neutral Flowering in Cannabis sativa L.

Hemp (Cannabis sativa L.) is a primarily short-day crop grown for grain, fiber, and secondary metabolites. Although terminal flowering of most non-domesticated hemp is regarded as photoperiod sensitive, limited germplasm is available for the development of day-neutral hemp. Day-neutral plants experience flower maturation independently of the short-day photoperiod cue which is typically triggered by photoperiods of less than 14 hours. The day-neutral trait is the subject of increased commercial interest for the purpose of breeding varieties with accelerated flowering time or cultivars that do not require labor- and cost-intensive light deprivation production systems. Genetic markers for the photoperiodic response would help breeders make early selection in the design of suitable cultivars for specific environments and cultivation calendars. Limited refereed information has been produced regarding the genetic regulation of photoperiodism in C. sativa. A population of 318 F2 individuals segregating for day-neutral flowering was developed, phenotyped, and genotyped. Genome-wide association analysis identified markers associated with the day-neutral trait indicating that a single recessive gene controlling photoperiod sensitivity is located within a large region of Chromosome 1. Flanking region sequence data of linked SNP markers was used in the development and validation of a TaqMan-based qPCR assay for the day-neutral trait. A genetic linkage map was produced, and QTL mapping identified two additional markers on Chromosome 1. Candidate genes, TARGET OF EARLY ACTIVATION TAGGED (TOE)/APETALA2 (AP2), and PSEUDO-RESPONSE REGULATOR 3 (PRR3), may work together to impact phase transition and photoperiodic flowering respectively and have key domains that are disrupted in day-neutral plants. The day-neutral flowering (a.k.a. "autoflower") phenotype is a valuable and understudied trait in Cannabis sativa The phenotype may result from a conserved linkage block of phase transition and photoperiodic flowering genes A segregating population was developed, genome wide analyses identified markers linked to the day-neutral trait Correlated markers were mapped to Chromosome 1 across a sizable region that appears to be largely non-recombining

plant biology↗

bioRxiv2023

PanKmer: k-mer based and reference-free pangenome analysis

SummaryPangenomes are replacing single reference genomes as the definitive representation of DNA sequence within a species or clade. Pangenome analysis predominantly leverages graph-based methods that require computationally intensive multiple genome alignments, do not scale to highly complex eukaryotic genomes, limit their scope to identifying structural variants (SVs), or incur bias by relying on a reference genome. Here, we present PanKmer, a toolkit designed for reference-free analysis of pangenome datasets consisting of dozens to thou-sands of individual genomes. PanKmer decomposes a set of input genomes into a table of observed k-mers and their presence-absence values in each genome. These are stored in an efficient k-mer index data format that encodes SNPs, INDELs, and SVs. It also includes functions for downstream analysis of the k-mer index, such as calculating sequence similarity statistics between individuals at whole-genome or local scales. For example, k-mers can be "anchored" in any individual genome to quantify sequence variability or conservation at a specific locus. This facilitates workflows with various biological applications, e.g. identifying cases of hybridization between plant species. PanKmer provides researchers with a valuable and convenient means to explore the full scope of genetic variation in a population, without reference bias. Availability and implementationPanKmer is implemented as a Python package with components written in Rust, released under a BSD license. The source code is available from the Python Package Index (PyPI) at https://pypi.org/project/pankmer/ as well as Gitlab at https://gitlab.com/salk-tm/pankmer. Full documentation is available at https://salk-tm.gitlab.io/pankmer/. Supplementary informationSupplementary data are available online

bioinformatics↗

bioRxiv2023

Generating high quality reference genomes from field collected specimens by optimizing preservation

Sample preservation challenges efforts to generate high-quality reference genomes or pangenomes for all 2 million+ plant and animal species. Here we show 95% ethanol protects against degradation for fish blood (22{degrees}C, [≤]6 weeks) and plant tissue (4{degrees}C, [≤]3 weeks). Using Nanopore, we assemble high quality reference genomes from three fish and two plant species (contig N50: 6.5-13.8Mb; BUSCO completeness: 94.4-99.2%; QV: 43.8 for M. esculenta).

genomics↗

bioRxiv2023

Optimization of molecular methods for detection and quantification of specific duckweed-bacteria interactions

Bacterial colonization dynamics of plants can differ between phylogenetically similar bacterial strains as well as in the context of complex bacterial communities. Quantitative studies that can resolve closely related bacteria within complex communities can lead to a better understanding of plant-microbe interactions. However, current methods lack the specificity to differentiate phylogenetically similar bacterial strains. In this study, we describe molecular strategies to study specific duckweed-bacteria interactions. We first systematically optimized a bead-beating protocol to co-isolate nucleic acids simultaneously from duckweed and bacteria. We then developed a generic fingerprinting assay to detect bacteria present in duckweed samples. To detect specific duckweed-bacteria interactions, we developed a genomics-based computational pipeline to generate bacterial strain-specific primers. These strain-specific primers differentiated bacterial strains from the same genus and enabled the detection of specific duckweed-bacteria interactions present in a community context. Moreover, we used these strain-specific primers to quantify the bacterial colonization of duckweed by normalization to a plant reference gene and revealed differences in colonization levels between strains from the same genus. Lastly, confocal microscopy of inoculated duckweed further supported our PCR results and showed bacterial colonization of the duckweed root-frond interface and root interior. The molecular methods introduced in this work should enable the tracking and quantification of specific plant-microbe interactions within plant-microbial communities.

ecology↗

bioRxiv2022

Genomics of turions from the Greater Duckweed reveal its pathways for dormancy and reemergence strategy

O_LIOver 15 families of aquatic plants are known to use a strategy of developmental switching upon environmental stress to produce dormant propagules called turions. However, few molecular details for turion biology have been elucidated due to the difficulties in isolating high-quality nucleic acids from this tissue. We successfully developed a new protocol to isolate high-quality transcripts and carried out RNA-seq analysis of mature turions from the Greater Duckweed Spirodela polyrhiza. Comparison of turion transcriptome to that of fronds, the actively growing leaf-like tissue, were carried out. C_LIO_LIBioinformatic analysis of high confidence, differentially expressed transcripts between frond and mature turion tissues revealed major pathways related to stress tolerance, starch and lipid metabolism, and dormancy that are mobilized to reprogram frond meristems for turion differentiation. C_LIO_LIWe identified the key genes that are likely to drive starch and lipid accumulation during turion formation, as well as in pathways for starch and lipid utilization upon turion germination. Comparison of genome-wide cytosine methylation levels also revealed evidence for epigenetic changes in the formation of turion tissues. C_LIO_LISimilarities between turions and seeds provided evidence that key regulators for seed maturation and germination have been retooled for their function in turion biology. C_LI

plant biology↗

bioRxiv2022

Time of day analysis over a field grown developmental time course in rice

Plants integrate time of day (TOD) information over an entire season to ensure optimal growth, flowering time, and grain fill. However, most TOD expression studies have focused on a limited number of combinations of daylength and temperature under laboratory conditions. Here we re-analyze an Oryza sativa (rice) expression study that followed TOD expression in the field over an entire growing season. We find that like Arabidopsis thaliana, almost all rice genes have a TOD specific expression over the developmental time course. As has been suggested in other grasses, thermocycles were a stronger cue for TOD expression than the photocycles over the growing season. All the core circadian clock genes display consistent TOD expression over the season with the interesting exception that the two grass paralogs of EARLY FLOWERING 3 (ELF3) display a distinct phasing based on the interaction between thermo- and photo-cycles. The dataset also revealed how specific pathways are modulated to distinct TOD over the season consistent with the changing biology. The data presented here provides a resource for researchers to study how TOD expression changes under natural conditions over a developmental time course, which will guide approaches to engineer more resilient and prolific crops.

plant biology↗

bioRxiv2022

Evolutionary gain and loss of a plant pattern-recognition receptor for HAMP recognition

Pattern recognition receptors (PRR) recognize distinct pathogen and herbivore-associated molecular patterns (PAMPs and HAMPs) and mediate activation of immune responses, but the evolution of new PRR sensing functions is not well understood. We employed comparative genomics and functional analysis to define evolutionary events leading to the sensing of the peptide HAMP inceptin (In11) by the PRR Inceptin Receptor (INR). Existing and de novo genome assemblies revealed that the presence of a functional INR gene corresponded with In11 response across 55 million years (my) of legume evolution, and that In11 recognition is unique to the clade of Phaseoloid legumes. The INR loci of certain Phaseoloid and non-Phaseoloid species also contain diverse INR-like homologues, suggesting that the evolution of INR receptor function [~]28 mya occurred after an ancestral gene insertion [~]32 mya. Functional analysis of chimeric and ancestrally reconstructed receptors revealed that specific AA differences in the C1 leucine-rich repeat (LRR) domain and C2 intervening motif likely mediated gain of In11 recognition. In summary, we present a conceptual model for the evolution of a defined PRR function based on patterns of INR variation in legumes.

plant biology↗

bioRxiv2022

Fish microbiomes 101: disentangling the rules governing marine fish mucosal microbiomes across 101 species

Fish are the most diverse and widely distributed vertebrates, yet little is known about the microbial ecology of fishes nor the biological and environmental factors that influence the fish microbiome. The microbiota from 101 species of Southern California marine fishes, spanning 22 orders, 55 families, and 83 genera representing ~25% of local marine fish diversity, was analyzed to identify patterns that explain microbial diversity patterns in a geographical subset of marine fish biodiversity. We compared fish microbiomes (gill, skin, midgut, and hindgut) using alpha, beta, and gamma diversity along with establishing a novel method to estimate microbial biomass (Qiime2 plugin katharoseq). For oceanic fishes from the neritic zone, host size and distance from shore were negatively associated with microbial biomass densities and diversity in the gills. Body site was the strongest driver for beta diversity with strong evidence of phylosymbiosis observed across the gill, skin, and hindgut, but not midgut. The majority of microbes from all fish body sites were of unknown origin but overall sea water generally contributes more microbes to fish mucus compared to marine sediment. In a meta-analysis of vertebrate hindguts (569 species), mammals had the highest gamma diversity when controlling for host species number while fishes had the highest percent of unique microbial taxa (92%). In fishes, the midgut, gill, and skin contains the majority of microbial diversity which collectively can be 5.5 times higher than the hindgut. The composite dataset will be useful to vertebrate microbiome researchers and fish biologists interested in microbial ecology with applications in aquaculture and fisheries management.

ecology↗

bioRxiv2021

Core circadian clock and light signalling genes brought into genetic linkage across the green lineage

The circadian clock ensures that biological processes are phased to the correct time of day. In plants the circadian clock is conserved at both the level of transcriptional networks as well as core genes. In the model plant Arabidopsis thaliana, the core circadian singleMYB (sMYB) genes CCA1 and RVE4 are in genetic linkage with the PSEUDO-RESPONSE REGULATOR (PRR) genes PRR9 and PRR7 respectively. Leveraging chromosome-resolved plant genomes and syntenic ortholog analysis it was possible to trace this genetic linkage back to the basal angiosperm Amborella and identify an additional evolutionarily conserved genetic linkage between PIF3 and PHYA. The LHY/CCA1-PRR5/9, RVE4/8-PRR3/7 and PIF3-PHYA genetic linkages emerged in the bryophyte lineage and progressively moved within several genes of each other across an array of higher plant families representing distinct whole genome duplication and fractionation events. Soybean maintains all but two genetic linkages, and expression analysis revealed the PIF3-PHYA linkage overlapping with the E4 maturity group locus was the only pair to robustly cycle with an evening phase in contrast to the sMYB-PRR morning and midday phase. While most monocots maintain the genetic linkages, they have been lost in the economically important grasses (Poaceae) such as maize where the genes have been fractionated to separate chromosomes and presence/absence variation results in the segregation of PRR7 paralogs across heterotic groups. The evolutionary conservation of the genetic linkage as well as its loss in the grasses provides new insight in the plant circadian clock, which has been a critical target of breeding and domestication. Summary SentenceThe genetic linkage of the core circadian clock components has evolutionary origins in bryophytes and sheds light on the current functioning and selection on the circadian clock in crops.

plant biology↗

bioRxiv2021

Chromosome-level genome assembly of a human fungal pathogen reveals synteny among geographically distinct species

Histoplasma capsulatum, a dimorphic fungal pathogen, is the most common cause of fungal respiratory infections in immunocompetent hosts. Histoplasma is endemic in the Ohio and Mississippi River Valleys in the United States and also distributed worldwide. Previous studies revealed at least eight clades, each specific to a geographic location: North American classes 1 and 2 (NAm 1 and NAm 2), Latin American groups A and B (LAm A and LAm B), Eurasian, Netherlands, Australian and African, and an additional distinct lineage (H81) comprised of Panamanian isolates. Previously assembled Histoplasma genomes are highly fragmented, with the highly repetitive G217B (NAm 2) strain, which has been used for most whole genome-scale transcriptome studies, assembled into over 250 contigs. In this study, we set out to fully assemble the repeat regions and characterize the large-scale genome architecture of Histoplasma species. We re-sequenced five Histoplasma strains (WU24 (NAm 1), G217B (NAm 2), H88 (African), G186AR (Panama), and G184AR (Panama)) using Oxford Nanopore Technologies long-read sequencing technology. Here we report chromosomal-level assemblies for all five strains, which exhibit extensive synteny among the geographically distant Histoplasma isolates. The new assemblies revealed that RYP2, a major regulator of morphology and virulence, is duplicated in G186AR. In addition, we mapped previously generated transcriptome datasets onto the newly assembled chromosomes. Our analyses revealed that the expression of transposons and transposon-embedded genes are upregulated in yeast phase compared to mycelial phase in G217B and H88 strains. This study provides an important resource for fungal researchers and further highlights the importance of chromosomal-level assemblies in analyzing high-throughput datasets. ImportanceHistoplasma species are dimorphic fungi causing significant morbidity and mortality worldwide. These fungi grow as mold in the soil and as budding yeast within the human host. Histoplasma can be isolated from soil in diverse regions, including North America, South America, Africa and Europe. Phylogenetically distinct species of Histoplasma have been isolated and sequenced. However, for the commonly used strains, genome assemblies have been fragmented, leading to underutilization of genome-scale data. This study provides chromosome-level assemblies of the commonly used Histoplasma strains using long-read sequencing technology. Comparative analysis of these genomes shows largely conserved gene order within the chromosomes. Mapping existing transcriptome data on these new assemblies reveals clustering of transcriptionally co-regulated genes. Results of this study highlight the importance of obtaining chromosome-level assemblies in understanding the biology of human fungal pathogens.

microbiology↗

bioRxiv2021

Contrasting a reference cranberry genome to a crop wild relative provides insights into adaptation, domestication, and breeding

Cranberry (Vaccinium macrocarpon) is a member of the Heath family (Ericaceae) and is a temperate low-growing woody perennial native to North America that is both economically important and has significant health benefits. While some native varieties are still grown today, breeding programs over the past 50 years have made significant contributions to improving disease resistance, fruit quality and yield. An initial genome sequence of an inbred line of the wild selection Ben Lear, which is parent to multiple breeding programs, provided insight into the gene repertoire as well as a platform for molecular breeding. Recent breeding efforts have focused on leveraging the circumboreal V. oxycoccos, which forms interspecific hybrids with V. macrocarpon, offering to bring in novel fruit chemistry and other desirable traits. Here we present an updated, chromosome-resolved V. macrocarpon reference genome, and compare it to a high-quality draft genome of V. oxycoccos. Leveraging the chromosome resolved cranberry reference genome, we confirmed that the Ericaceae has undergone two whole genome duplications that are shared with blueberry and rhododendron. Leveraging resequencing data for Ben Lear inbred lines, as well as several wild and elite selections, we identified common regions that are targets of improvement. These same syntenic regions in V. oxycoccos, were identified and represent environmental response and plant architecture genes. These data provide insight into early genomic selection in the domestication of a native North American berry crop.

genomics↗

bioRxiv2021

Large structural variations in the haplotype-resolved African cassava genome.

Cassava (Manihot esculenta Crantz, 2n=36) is a global food security crop. Cassava has a highly heterozygous genome, high genetic load, and genotype-dependent asynchronous flowering. It is typically propagated by stem cuttings and any genetic variation between haplotypes, including large structural variations, is preserved by such clonal propagation. Traditional genome assembly approaches generate a collapsed haplotype representation of the genome. In highly heterozygous plants, this results in artifacts and an oversimplification of heterozygous regions. We used a combination of Pacific Biosciences (PacBio), Illumina, and Hi-C to resolve each haplotype of the genome of a farmer-preferred cassava line, TME7 (Oko-iyawo). PacBio reads were assembled using the FALCON suite. Phase switch errors were corrected using FALCON-Phase and Hi-C read data. The ultra-long-range information from Hi-C sequencing was also used for scaffolding. Comparison of the two phases revealed more than 5,000 large haplotype-specific structural variants affecting over 8 Mb, including insertions and deletions spanning thousands of base pairs. The potential of these variants to affect allele specific expression was further explored. RNA-seq data from 11 different tissue types were mapped against the scaffolded haploid assembly and gene expression data are incorporated into our existing easy-to-use web-based interface to facilitate use by the broader plant science community. These two assemblies provide an excellent means to study the effects of heterozygosity, haplotype-specific structural variation, gene hemizygosity, and allele specific gene expression contributing to important agricultural traits and further our understanding of the genetics and domestication of cassava. Significance statementThe cassava varieties grown by subsistence farmers in Africa largely differ from the inbred reference genome due to their highly heterozygous nature. We used multiple sequencing technologies to assemble and resolve both haplotypes in TME7, a farmer-preferred cassava line, enabling us to study the considerable haplotypic structural variation in this line.

genomics↗

bioRxiv2021

Underwater CAM photosynthesis elucidated by Isoetes genome

To conserve water in arid environments, numerous plant lineages have independently evolved Crassulacean Acid Metabolism (CAM). Interestingly, Isoetes, an aquatic lycophyte, can also perform CAM as an adaptation to low CO2 availability underwater. However, little is known about the evolution of CAM in aquatic plants and the lack of genomic data has hindered comparison between aquatic and terrestrial CAM. Here, we investigated the underwater CAM in Isoetes taiwanensis by generating a high-quality genome assembly and RNA-seq time course. Despite broad similarities between CAM in Isoetes and terrestrial angiosperms, we identified several key differences. Notably, for carboxylation of PEP, Isoetes recruited the lesser-known "bacterial-type" PEPC, along with the "plant-type" exclusively used in other terrestrial CAM and C4 plants. Furthermore, we found that circadian control of key CAM pathway genes has diverged considerably in Isoetes relative to flowering plants. This suggests the existence of more evolutionary paths to CAM than previously recognized.

plant biology↗