Evolutionary divergence of subgenomes in common carp provides insights into speciation and allopolyploid success.

Hybridization and polyploidization have made great contributions to speciation, heterosis, and agricultural production within plants, but there is still limited understanding and utilization in animals. Subgenome structure and expression reorganization and cooperation post hybridization and polyploidization are essential for speciation and allopolyploid success. However, the mechanisms have not yet been comprehensively assessed in animals. Here, we produced a high-fidelity reference genome sequence for common carp, a typical allotetraploid fish species cultured worldwide. This genome enabled in-depth analysis of the evolution of subgenome architecture and expression responses. Most genes were expressed with subgenome biases, with a trend of transition from the expression of subgenome A during the early stages to that of subgenome B during the late stages of embryonic development. While subgenome A evolved more rapidly, subgenome B contributed to a greater level of expression during development and under stressful conditions. Stable dominant patterns for homoeologous gene pairs both during development and under thermal stress suggest a potential fixed heterosis in the allotetraploid genome. Preferentially expressing either copy of a homoeologous gene at higher levels to confer development and response to stress indicates the dominant effect of heterosis. The plasticity of subgenomes and their shifting of dominant expression during early development, and in response to stressful conditions, provide novel insights into the molecular basis of the successful speciation, evolution, and heterosis of the allotetraploid common carp.

Intersectional Hybrids between Darrow's Blueberry (V. darrowii Camp) and Lingonberry (V. vitis-idaea L.).

An initial cross of V. darrowii 'Johnblue' (Darrow's blueberry) × V. vitis-idaea 'Red Sunset' (lingonberry) produced more than 30 true intersectional diploid hybrids as confirmed by molecular markers. The most vigorous of these hybrids was extensively evaluated. This hybrid, US 2535-A, was floriferous and morphologically intermediate to the respective parents. Examination of pollen suggested low male fertility. Numerous crosses using the hybrid as a female reflected similarly low fertility and potential crossing barriers. Stylar examination suggested blockage of pollen tube growth in self-pollinations and significantly retarded growth in backcross pollinations. Nonetheless, two confirmed hybrid offspring were produced using the F1 hybrid as a female in crosses with V. vitis-idaea and V. darrowii, respectively. In a second set of crosses utilizing additional V. darrowii and V. vitis-idaea genotypes, another 23 verified hybrids in seven parental combinations were produced. Hybrids such as the ones presented offer the potential for generating de novo interspecific fruit types in blueberry and/or broadening the adaptation of lingonberry.

Subgenomic T cell receptor alpha and delta (TRA/TRD) loci in common carp.

In jawed vertebrates, the T cell receptor alpha (TRA) and delta (TRD) genes, which encode the TRα and TRδ chains, respectively, are located as a nested structure on a single chromosome. To date, no animal has been reported to harbor multiple TRA/TRD loci on different chromosomes. Therefore, herein, we describe the first full annotation of the TRA/TRD genomic regions of common carp, an allo-tetraploid fish species that experiences cyprinid-specific whole-genome duplication (WGD) in evolution. Fine genomic maps of TRA/TRD genomic regions 1 and 2, on LG30 and LG22, respectively, were constructed using the annotations of complete sets of TRA and TRD genes, including TRA/TRD variable (V), TRA junction (J), and constant (C), TRD diversity (D), and the J and C genes. The structure and synteny of the TRA/TRD genomic regions were highly conserved in zebrafish, indicating that these regions are on individual chromosomes. Furthermore, analysis of the variable regions of the TRA and TRD genes in a monoclonal T cell line revealed that both subgenomic regions 1 and 2 were indeed rearranged. Although carp TRAV and TRDV genes were phylogenetically divided into different lineages, they were mixed and organized into the TRA/TRD V gene clusters on the genome, similar to that in other vertebrates. Notably, 285 potential TRA/TRD V genes were detected in the TRA/TRD genomic regions, which is the most abundant number of genes in vertebrates and approximately two-fold that in zebrafish. The recombination signal sequences (RSSs) at the end of each V gene differed between TRAV and TRDV, suggesting that RSS variations might separate each V gene into a TRα or TRδ chain. This study is the first to describe subgenomic TRA/TRD loci in animals. Our findings provide fundamental insights to elucidate the impact of WGD on the evolution of immune repertoire.

Pectin demethylation-mediated cell wall Na+ retention positively regulates salt stress tolerance in oilseed rape.

KEY MESSAGE: Integrated phenomics, ionomics, genomics, transcriptomics, and functional analyses present novel insights into the role of pectin demethylation-mediated cell wall Na+ retention in positively regulating salt tolerance in oilseed rape. Genetic variations in salt stress tolerance identified in rapeseed genotypes highlight the complicated regulatory mechanisms. Westar is ubiquitously used as a transgenic receptor cultivar, while ZS11 is widely grown as a high-production and good-quality cultivar. In this study, Westar was found to outperform ZS11 under salt stress. Through cell component isolation, non-invasive micro-test, X-ray energy spectrum analysis, and ionomic profile characterization, pectin demethylation-mediated cell wall Na+ retention was proposed to be a major regulator responsible for differential salt tolerance between Westar and ZS11. Integrated analyses of genome-wide DNA variations, differential expression profiling, and gene co-expression networks identified BnaC9.PME47, encoding a pectin methylesterase, as a positive regulator conferring salt tolerance in rapeseed. BnaC9.PME47, located in two reported QTL regions for salt tolerance, was strongly induced by salt stress and localized on the cell wall. Natural variation of the promoter regions conferred higher expression of BnaC9.PME47 in Westar than in several salt-sensitive rapeseed genotypes. Loss of function of AtPME47 resulted in the hypersensitivity of Arabidopsis plants to salt stress. The integrated multiomics analyses revealed novel insights into pectin demethylation-mediated cell wall Na+ retention in regulating differential salt tolerance in allotetraploid rapeseed genotypes. Furthermore, these analyses have provided key information regarding the rapid dissection of quantitative trait genes responsible for nutrient stress tolerance in plant species with complex genomes.

Genomic and cytogenetic analyses reveal satellite repeat signature in allotetraploid okra (Abelmoschus esculentus).

BACKGROUND: Satellite repeats are one of the most rapidly evolving components in eukaryotic genomes and play vital roles in genome regulation, genome evolution, and speciation. As a consequence, the composition, abundance and chromosome distribution of satellite repeats often exhibit variability across various species, genome, and even individual chromosomes. However, we know little about the satellite repeat evolution in allopolyploid genomes. RESULTS: In this study, we investigated the satellite repeat signature in five okra (Abelmoschus esculentus) accessions using genomic and cytogenetic methods. In each of the five accessions, we identified eight satellite repeats, which exhibited a significant level of intraspecific conservation. Through fluorescence in situ hybridization (FISH) experiments, we observed that the satellite repeats generated multiple signals and exhibited variations in copy number across chromosomes. Intriguingly, we found that five satellite repeats were interspersed with centromeric retrotransposons, signifying their involvement in centromeric satellite repeat identity. We confirmed subgenome-biased amplification patterns of these satellite repeats through existing genome assemblies or dual-color FISH, indicating their distinct dynamic evolution in the allotetraploid okra subgenome. Moreover, we observed the presence of multiple chromosomes harboring the 35 S rDNA loci, alongside another chromosomal pair carrying the 5 S rDNA loci in okra using FISH assay. Remarkably, the intensity of 35 S rDNA hybridization signals varied among chromosomes, with the signals predominantly localized within regions of relatively weak DAPI staining, associated with GC-rich heterochromatin regions. Finally, we observed a similar localization pattern between 35 S rDNA and three satellite repeats with high GC content and confirmed their origin in the intergenic spacer region of the 35 S rDNA. CONCLUSIONS: Our findings uncover a unique satellite repeat signature in the allotetraploid okra, contributing to our understanding of the composition, abundance, and chromosomal distribution of satellite repeats in allopolyploid genomes, further enriching our understanding of their evolutionary dynamics in complex allopolyploid genomes.

The complete chloroplast genome of the neo-allotetraploid fern, Asplenium pseudocapillipes S.H.Park et al. (Aspleniaceae).

This study analyzed the complete plastome sequence of the neo-allotetraploid Asplenium pseudocapillipes S.H.Park et al. Asplenium pseudocapillipes has a typical circular plastome that comprises 157,242 bp with a large single copy (84,105 bp), a small single copy (21,503 bp), and two inverted repeats (IRs; 25,817 bp). The complete sequence comprises 127 genes, including 87 protein-coding genes (CDSs), eight ribosomal RNAs (rRNAs), 31 transfer RNAs (tRNAs), and one pseudogene. Among these genes, five CDSs, four rRNAs, and five tRNAs are duplicated in IRs. The guanine-cytosine content of the genome was 41.5%. The enlarged noncoding regions by Mobile Open Reading Frames in Fern Organelles were found once in other Asplenium species and twice in A. pseudocapillipes. Phylogenetic analysis based on 83 coding gene sequences revealed that A. pseudocapillipes is embedded in the A. varians subclade along with its progenitors.

Telomere-to-telomere genome of the allotetraploid legume Sesbania cannabina reveals transposon-driven subgenome divergence and mechanisms of alkaline stress tolerance.

Alkaline soils pose an increasing problem for agriculture worldwide, but using stress-tolerant plants as green manure can improve marginal land. Here, we show that the legume Sesbania cannabina is very tolerant to alkaline conditions and, when used as a green manure, substantially improves alkaline soil. To understand genome evolution and the mechanisms of stress tolerance in this allotetraploid legume, we generated the first telomere-to-telomere genome assembly of S. cannabina spanning ∼2,087 Mb. The assembly included all centromeric regions, which contain centromeric satellite repeats, and complete chromosome ends with telomeric characteristics. Further genome analysis distinguished A and B subgenomes, which diverged approximately 7.9 million years ago. Comparative genomic analysis revealed that the chromosome homoeologs underwent large-scale inversion events (>10 Mb) and a significant, transposon-driven size expansion of the chromosome 5A homoeolog. We further identified four specific alkali-induced phosphate transporter genes in S. cannabina; these may function in alkali tolerance by relieving the deficiency in available phosphorus in alkaline soil. Our work highlights the significance of S. cannabina as a green tool to improve marginal lands and sheds light on subgenome evolution and adaptation to alkaline soils.

Genome-wide identification of the cation/proton antiporter (CPA) gene family and functional characterization of the key member BnaA05.NHX2 in allotetraploid rapeseed.

Rapeseed (Brassica napus L.) is susceptible to nutrient stresses during growth and development; however, the CPA (cation proton antiporter) family genes have not been identified in B. napus and their biological functions remain unclear. This study was aimed to identify the molecular characteristics of rapeseed CPAs and their transcriptional responses to multiple nutrient stresses. Through bioinformatics analysis, 117 BnaCPAs, consisting of three subfamilies: Na+/H+ antiporter (NHX), K+ efflux antiporter (KEA), and cation/H+ antiporter (CHX), were identified in the rapeseed genome. Transcriptomic profiling showed that BnaCPAs, particularly BnaNHXs, were transcriptionally responsive to diverse nutrient stresses, including Cd toxicity, K starvation, salt stress, NH4+ toxicity, and low Pi. We found that the salt tolerance of the transgenic rapeseed lines overexpressing BnaA05.NHX2 was significantly higher than that of wild type. Subcellular localization showed that BnaA05.NHX2 was localized on the tonoplast, and TEM combined with X-ray energy spectrum analysis revealed that the vacuolar Na+ concentrations of the BnaA05.NHX2-overexpressing rapeseed plants were significantly higher than those of wild type. The findings of this study will provide insights into the complexity of the BnaCPA family and a valuable resource to explore the in-depth functions of CPAs in B. napus.

The evolution history of an allotetraploid mangrove tree analysed with a new tool Allo4D.

Mangrove species are broadly classified as true mangroves and mangrove associates. The latter are amphibious plants that can survive in the intertidal zone and reproduce naturally in terrestrial environments. Their widespread distribution and extensive adaptability make them ideal research materials for exploring adaptive evolution. In this study, we de novo assembled two genomes of mangrove associates (the allotetraploid Barringtonia racemosa (2n = 4x = 52) and diploid Barringtonia asiatica (2n = 2x = 26)) to investigate the role of allopolyploidy in the evolutionary history of mangrove species. We developed a new allotetraploid-dividing tool Allo4D to distinguish between allotetraploid scaffold-scale subgenomes and verified its accuracy and reliability using real and simulated data. According to the two subgenomes of allotetraploid B. racemosa divided using Allo4D, the allopolyploidization event was estimated to have occurred approximately one million years ago (Mya). We found that B. racemosa, B. asiatica, and Diospyros lotus shared a whole genome duplication (WGD) event during the K-Pg (Cretaceous-Paleozoic) period. K-Pg WGD and recent allopolyploidization events contributed to the speciation of B. racemosa and its adaptation to coastal habitats. We found that genes in the glucosinolates (GSLs) pathway, an essential pathway in response to various biotic and abiotic stresses, expanded rapidly in B. racemosa during polyploidization. In summary, this study provides a typical example of the adaptation of allopolyploid plants to extreme environmental conditions. The newly developed tool, Allo4D, can effectively divide allotetraploid subgenomes and explore the evolutionary history of polyploid plants, especially for species whose ancestors are unknown or extinct.

Development of an SNP marker set for marker-assisted backcrossing using genotyping-by-sequencing in tetraploid perilla.

High-quality molecular markers are essential for marker-assisted selection to accelerate breeding progress. Compared with diploid species, recently diverged polyploid crop species tend to have highly similar homeologous subgenomes, which is expected to limit the development of broadly applicable locus-specific single-nucleotide polymorphism (SNP) assays. Furthermore, it is particularly challenging to make genome-wide marker sets for species that lack a reference genome. Here, we report the development of a genome-wide set of kompetitive allele specific PCR (KASP) markers for marker-assisted recurrent selection (MARS) in the tetraploid minor crop perilla. To find locus-specific SNP markers across the perilla genome, we used genotyping-by-sequencing (GBS) to construct linkage maps of two F2 populations. The two resulting high-resolution linkage maps comprised 2326 and 2454 SNP markers that spanned a total genetic distance of 2133 cM across 16 linkage groups and 2169 cM across 21 linkage groups, respectively. We then obtained a final genetic map consisting of 22 linkage groups with 1123 common markers from the two genetic maps. We selected 96 genome-wide markers for MARS and confirmed the accuracy of markers in the two F2 populations using a high-throughput Fluidigm system. We confirmed that 91.8% of the SNP genotyping results from the Fluidigm assay were the same as the results obtained through GBS. These results provide a foundation for marker-assisted backcrossing and the development of new varieties of perilla.

Haplotype-Resolved, Chromosome-Level Assembly of White Clover (Trifolium repens L., Fabaceae).

White clover (Trifolium repens L.; Fabaceae) is an important forage and cover crop in agricultural pastures around the world and is increasingly used in evolutionary ecology and genetics to understand the genetic basis of adaptation. Historically, improvements in white clover breeding practices and assessments of genetic variation in nature have been hampered by a lack of high-quality genomic resources for this species, owing in part to its high heterozygosity and allotetraploid hybrid origin. Here, we use PacBio HiFi and chromosome conformation capture (Omni-C) technologies to generate a chromosome-level, haplotype-resolved genome assembly for white clover totaling 998 Mbp (scaffold N50 = 59.3 Mbp) and 1 Gbp (scaffold N50 = 58.6 Mbp) for haplotypes 1 and 2, respectively, with each haplotype arranged into 16 chromosomes (8 per subgenome). We additionally provide a functionally annotated haploid mapping assembly (968 Mbp, scaffold N50 = 59.9 Mbp), which drastically improves on the existing reference assembly in both contiguity and assembly accuracy. We annotated 78,174 protein-coding genes, resulting in protein BUSCO completeness scores of 99.6% and 99.3% against the embryophyta_odb10 and fabales_odb10 lineage datasets, respectively.

Histone Maps in Gossypium darwinii Reveal Epigenetic Regulation Drives Subgenome Divergence and Cotton Domestication.

The functional annotation of genomes, including chromatin modifications, is essential to understand the intricate architecture of chromatin and the consequential gene regulation. However, such an annotation remains limited for cotton genomes. Here, we conducted chromatin profiling in a wild allotetraploid cotton Gossypium darwinii (AD genome) by integrating the data of histone modification, transcriptome, and chromatin accessibility. We revealed that the A subgenome showed a higher level of active histone marks and lower level of repressive histone marks than the D subgenome, which was consistent with the expression bias between the two subgenomes. We show that the bias in transcription and histone modification between the A and D subgenomes may be caused by genes unique to the subgenome but not by homoeologous genes. Moreover, we integrate histone marks and open chromatin to define six chromatin states (S1-S6) across the cotton genome, which index different genomic elements including genes, promoters, and transposons, implying distinct biological functions. In comparison to the domesticated cotton species, we observed that 23.2% of genes in the genome exhibit a transition from one chromatin state to another at their promoter. Strikingly, the S2 (devoid of epigenetic marks) to S3 (enriched for the mark of open chromatin) was the largest transition group. These transitions occurred simultaneously with changes in gene expression, which were significantly associated with several domesticated traits in cotton. Collectively, our study provides a useful epigenetic resource for research on allopolyploid plants. The domestication-induced chromatin dynamics and associated genes identified here will aid epigenetic engineering, improving polyploid crops.

Genome-wide identification of Brassicaceae histone modification genes and their responses to abiotic stresses in allotetraploid rapeseed.

BACKGROUND: Histone modification is an important epigenetic regulatory mechanism and essential for stress adaptation in plants. However, systematic analysis of histone modification genes (HMs) in Brassicaceae species is lacking, and their roles in response to abiotic stress have not yet been identified. RESULTS: In this study, we identified 102 AtHMs, 280 BnaHMs, 251 BcHMs, 251 BjHMs, 144 BnHMs, 155 BoHMs, 137 BrHMs, 122 CrHMs, and 356 CsHMs in nine Brassicaceae species, respectively. Their chromosomal locations, protein/gene structures, phylogenetic trees, and syntenies were determined. Specific domains were identified in several Brassicaceae HMs, indicating an association with diverse functions. Syntenic analysis showed that the expansion of Brassicaceae HMs may be due to segmental and whole-genome duplications. Nine key BnaHMs in allotetraploid rapeseed may be responsible for ammonium, salt, boron, cadmium, nitrate, and potassium stress based on co-expression network analysis. According to weighted gene co-expression network analysis (WGCNA), 12 BnaHMs were associated with stress adaptation. Among the above genes, BnaPRMT11 simultaneously responded to four different stresses based on differential expression analysis, while BnaSDG46, BnaHDT10, and BnaHDA1 participated in five stresses. BnaSDG46 was also involved in four different stresses based on WGCNA, while BnaSDG10 and BnaJMJ58 were differentially expressed in response to six different stresses. In summary, six candidate genes for stress resistance (BnaPRMT11, BnaSDG46, BnaSDG10, BnaJMJ58, BnaHDT10, and BnaHDA1) were identified. CONCLUSIONS: Taken together, these findings help clarify the biological roles of Brassicaceae HMs. The identified candidate genes provide an important reference for the potential development of stress-tolerant oilseed plants.

Marker-assisted introgression of wild chromosome segments conferring resistance to fungal foliar diseases into peanut (Arachis hypogaea L.).

Introduction: Fungal foliar diseases can severely affect the productivity of the peanut crop worldwide. Late leaf spot is the most frequent disease and a major problem of the crop in Brazil and many other tropical countries. Only partial resistance to fungal diseases has been found in cultivated peanut, but high resistances have been described on the secondary gene pool. Methods: To overcome the known compatibility barriers for the use of wild species in peanut breeding programs, we used an induced allotetraploid (Arachis stenosperma × A. magna)4x, as a donor parent, in a successive backcrossing scheme with the high-yielding Brazilian cultivar IAC OL 4. We used microsatellite markers associated with late leaf spot and rust resistance for foreground selection and high-throughput SNP genotyping for background selection. Results: With these tools, we developed agronomically adapted lines with high cultivated genome recovery, high-yield potential, and wild chromosome segments from both A. stenosperma and A. magna conferring high resistance to late leaf spot and rust. These segments include the four previously identified as having QTLs (quantitative trait loci) for resistance to both diseases, which could be confirmed here, and at least four additional QTLs identified by using mapping populations on four generations. Discussion: The introgression germplasm developed here will extend the useful genetic diversity of the primary gene pool by providing novel wild resistance genes against these two destructive peanut diseases.

Genome-wide analysis of R2R3-MYB genes in cultivated peanut (Arachis hypogaea L.): Gene duplications, functional conservation, and diversification.

The cultivated Peanut (Arachis hypogaea L.), an important oilseed and edible legume, are widely grown worldwide. The R2R3-MYB transcription factor, one of the largest gene families in plants, is involved in various plant developmental processes and responds to multiple stresses. In this study we identified 196 typical R2R3-MYB genes in the genome of cultivated peanut. Comparative phylogenetic analysis with Arabidopsis divided them into 48 subgroups. The motif composition and gene structure independently supported the subgroup delineation. Collinearity analysis indicated polyploidization, tandem, and segmental duplication were the main driver of the R2R3-MYB gene amplification in peanut. Homologous gene pairs between the two subgroups showed tissue specific biased expression. In addition, a total of 90 R2R3-MYB genes showed significant differential expression levels in response to waterlogging stress. Furthermore, we identified an SNP located in the third exon region of AdMYB03-18 (AhMYB033) by association analysis, and the three haplotypes of the SNP were significantly correlated with total branch number (TBN), pod length (PL) and root-shoot ratio (RS ratio), respectively, revealing the potential function of AdMYB03-18 (AhMYB033) in improving peanut yield. Together, these studies provide evidence for functional diversity in the R2R3-MYB genes and will contribute to understanding the function of R2R3-MYB genes in peanut.

Whole-genome sequence of synthetically derived Brassica napus inbred cultivar Da-Ae.

Brassica napus, a globally important oilseed crop, is an allotetraploid hybrid species with two subgenomes originating from Brassica rapa and Brassica oleracea. The presence of two highly similar subgenomes has made the assembly of a complete draft genome challenging and has also resulted in natural homoeologous exchanges between the genomes, resulting in variations in gene copy number, which further complicates assigning sequences to correct chromosomes. Despite these challenges, high-quality draft genomes of this species have been released. Using third generation sequencing and assembly technologies, we generated a new genome assembly for the synthetic B. napus cultivar Da-Ae. Through the use of long reads, linked-reads, and Hi-C proximity data, we assembled a new draft genome that provides a high-quality reference genome of a synthetic B. napus. In addition, we identified potential hotspots of homoeologous exchange between subgenomes within Da-Ae, based on their presence in other independently derived lines. The occurrence of these hotspots may provide insight into the genetic rearrangements required for B. napus to be viable following the hybridization of B. rapa and B. oleracea.

The wild allotetraploid sesame genome provides novel insights into evolution and lignan biosynthesis.

INTRODUCTION: The wild tetraploid sesame (Sesamum schinzianum), an ancestral relative of diploid cultivated sesame, grows in the tropical desert of the African Plateau. As a valuable seed resource, wild sesame has several advantageous traits, such as strong environmental adaptability and an extremely high content of sesamolin in its seeds. High-quality genome assembly is essential for a detailed understanding of genome structure, genome evolution and crop improvement. OBJECTIVES: Here, we generated two high-quality chromosome-scale genomes from S. schinzianum and a cultivated diploid elite sesame (Sesamum indicum L.) to investigate the potential genetic basis underlying these traits of wild sesame. METHODS: The long-read data from PacBio Sequel II platform and high-throughput chromosome conformation capture (Hi-C) data were used to construct high-quality sesame genome. Then dissecting the molecular mechanisms of sesame evolution and lignan biosynthesis through comparative genomics and transcriptomics. RESULTS: We found evidence of divergent evolution that involved differences in the number, sequence and expression level of homologous genes between the two sets of subgenomes from allotetraploids in S. schinzianum, all of which might be driven by subfunctionalization after polyploidization. Furthermore, it was found that a great number of genes involved in the stress response have undergone positive selection and resulted from gene family expansion in the wild sesame genome compared with the cultivated sesame genome, which, overall, is associated with adaptative evolution to the environment. We hypothesized that the sole functional member CYP92B14 (SscC22g35272) could be associated with high content of sesamolin in wild sesame seeds. CONCLUSION: This study provides high-quality wild allotetraploid sesame and cultivated sesame genomes, reveals evolutionary features of the allotetraploid genome and provides novel insights into lignan synthesis pathways. Meanwhile, the wild sesame genome will be an important resource to conduct comparative genomic and evolutionary studies and plant improvement programmes.

Down, then up: non-parallel genome size changes and a descending chromosome series in a recent radiation of the Australian allotetraploid plant species, Nicotiana section Suaveolentes (Solanaceae).

BACKGROUND AND AIMS: The extent to which genome size and chromosome numbers evolve in concert is little understood, particularly after polyploidy (whole-genome duplication), when a genome returns to a diploid-like condition (diploidization). We study this phenomenon in 46 species of allotetraploid Nicotiana section Suaveolentes (Solanaceae), which formed <6 million years ago and radiated in the arid centre of Australia. METHODS: We analysed newly assessed genome sizes and chromosome numbers within the context of a restriction site-associated nuclear DNA (RADseq) phylogenetic framework. KEY RESULTS: RADseq generated a well-supported phylogenetic tree, in which multiple accessions from each species formed unique genetic clusters. Chromosome numbers and genome sizes vary from n = 2x = 15 to 24 and 2.7 to 5.8 pg/1C nucleus, respectively. Decreases in both genome size and chromosome number occur, although neither consistently nor in parallel. Species with the lowest chromosome numbers (n = 15-18) do not possess the smallest genome sizes and, although N. heterantha has retained the ancestral chromosome complement, n = 2x = 24, it nonetheless has the smallest genome size, even smaller than that of the modern representatives of ancestral diploids. CONCLUSIONS: The results indicate that decreases in genome size and chromosome number occur in parallel down to a chromosome number threshold, n = 20, below which genome size increases, a phenomenon potentially explained by decreasing rates of recombination over fewer chromosomes. We hypothesize that, more generally in plants, major decreases in genome size post-polyploidization take place while chromosome numbers are still high because in these stages elimination of retrotransposons and other repetitive elements is more efficient. Once such major genome size change has been accomplished, then dysploid chromosome reductions take place to reorganize these smaller genomes, producing species with small genomes and low chromosome numbers such as those observed in many annual angiosperms, including Arabidopsis.

Transcriptomic analysis of succulent stem development of Chinese kale (Brassica oleracea var. alboglabra Bailey) and its synthetic allotetraploid via RNA sequencing.

Chinese kale (Brassica oleracea var. alboglabra Bailey, CC) is a succulent stem vegetable in the Brassica family. Its allotetraploid (AACC) vegetable germplasm, which was synthesized via distant hybridization with the colloquially named 'yellow turnip' (B. rapa L. ssp. rapifera Matzg., AA), has a swelling stem similar to CC. To address the molecular mechanism of stem development for CC and AACC, RNA sequencing (RNA-seq) was used to investigate transcriptional regulation of their stem development at three key stages including 28 days, 42 days and the bolting stage (BS) after sowing. As a result, 32,642, 32,665, 33,816, 32,147, 32,293 and 32,275 genes were identified in six corresponding cDNA libraries. Among them, 25,459 genes were co-expressed, while 7,183, 7,206, 8,357, 6,688, 6,834 and 6,814 genes were specifically expressed. Additionally, a total of 29,222 differentially expressed genes (DEGs) were found for functional enrichment as well as many genes involved in plant hormones including gibberellin (GA), abscisic acid (ABA), cytokinin (CTK) and auxin (AUX). Based on gene expression consistency between CC and AACC, the gene families including DELLA, GID, PYR/PYL, PP2C, A-ARR and AUX/IAA might be related to stem development. Among these, eight genes including Bo00834s040, Bo5g093140, Bo6g086770, Bo9g070200, Bo7g116570, Bo3g054410, Bo7g093470 and Bo5g136600 may play important roles in stem development based on their remarkable expression levels as confirmed by qRT-PCR. These findings provide a new theoretical basis for understanding the molecular mechanism of stem development in Brassica vegetable stem breeding.

Asplenium pseudocapillipes (Aspleniaceae), a New Fern Species from South Korea.

A new allotetraploid species of the genus Asplenium, A. pseudocapillipes, originated from the hybridization between A. capillipes and A. tenuicaule, has been newly discovered in two limestone areas of South Korea. A molecular phylogenetic analysis using one chloroplast region (rbcL) and three single- or low-copy nuclear regions (AK1, gapCp, pgiC) and a cytological analysis, including genome size measurements, were conducted to characterize this new species. From these results, the maternal origin of A. pseudocapillipes was confirmed to be A. capillipes, which has never been reported in Korea. All three nuclear data showed that this new species had genotypes of both A. capillipes and A. tenuicaule. The quantitative characteristics of the leaves showed values intermediate between the two parental species. The absence of gemma accorded with its paternal origin from A. tenuicaule, and 32 spores per sporangium accorded with its maternal origin from A. capillipes. Although A. pseudocapillipes has 32 spores per sporangium, it is considered to be a sexually reproducing, not an apomitic, fern.