A chromosomal-scale genome assembly of modern cultivated hybrid sugarcane provides insights into origination and evolution.

Sugarcane is a vital crop with significant economic and industrial value. However, the cultivated sugarcane's ultra-complex genome still needs to be resolved due to its high ploidy and extensive recombination between the two subgenomes. Here, we generate a chromosomal-scale, haplotype-resolved genome assembly for a hybrid sugarcane cultivar ZZ1. This assembly contains 10.4 Gb genomic sequences and 68,509 annotated genes with defined alleles in two sub-genomes distributed in 99 original and 15 recombined chromosomes. RNA-seq data analysis shows that sugar accumulation-associated gene families have been primarily expanded from the ZZSO subgenome. However, genes responding to pokkah boeng disease susceptibility have been derived dominantly from the ZZSS subgenome. The region harboring the possible smut resistance genes has expanded significantly. Among them, the expansion of WAK and FLS2 families is proposed to have occurred during the breeding of ZZ1. Our findings provide insights into the complex genome of hybrid sugarcane cultivars and pave the way for future genomics and molecular breeding studies in sugarcane.

MIKE: an ultrafast, assembly-, and alignment-free approach for phylogenetic tree construction.

MOTIVATION: Constructing a phylogenetic tree requires calculating the evolutionary distance between samples or species via large-scale resequencing data, a process that is both time-consuming and computationally demanding. Striking the right balance between accuracy and efficiency is a significant challenge. RESULTS: To address this, we introduce a new algorithm, MIKE (MinHash-based k-mer algorithm). This algorithm is designed for the swift calculation of the Jaccard coefficient directly from raw sequencing reads and enables the construction of phylogenetic trees based on the resultant Jaccard coefficient. Simulation results highlight the superior speed of MIKE compared to existing state-of-the-art methods. We used MIKE to reconstruct a phylogenetic tree, incorporating 238 yeast, 303 Zea, 141 Ficus, 67 Oryza, and 43 Saccharum spontaneum samples. MIKE demonstrated accurate performance across varying evolutionary scales, reproductive modes, and ploidy levels, proving itself as a powerful tool for phylogenetic tree construction. AVAILABILITY AND IMPLEMENTATION: MIKE is publicly available on Github at https://github.com/Argonum-Clever2/mike.git.

Gene mining and genomics-assisted breeding empowered by the pangenome of tea plant Camellia sinensis.

Tea is one of the world's oldest crops and is cultivated to produce beverages with various flavours. Despite advances in sequencing technologies, the genetic mechanisms underlying key agronomic traits of tea remain unclear. In this study, we present a high-quality pangenome of 22 elite cultivars, representing broad genetic diversity in the species. Our analysis reveals that a recent long terminal repeat burst contributed nearly 20% of gene copies, introducing functional genetic variants that affect phenotypes such as leaf colour. Our graphical pangenome improves the efficiency of genome-wide association studies and allows the identification of key genes controlling bud flush timing. We also identified strong correlations between allelic variants and flavour-related chemistries. These findings deepen our understanding of the genetic basis of tea quality and provide valuable genomic resources to facilitate its genomics-assisted breeding.

5mC DNA methylation modification-mediated regulation in tissue functional differentiation and important flavor substance synthesis of tea plant (Camellia sinensis L.).

In plants, 5mC DNA methylation is an important and conserved epistatic mark involving genomic stability, gene transcriptional regulation, developmental regulation, abiotic stress response, metabolite synthesis, etc. However, the roles of 5mC DNA methylation modification (5mC methylation) in tea plant growth and development (in pre-harvest processing) and flavor substance synthesis in pre- and post-harvest processing are unknown. We therefore conducted a comprehensive methylation analysis of four key pre-harvest tissues (root, leaf, flower, and fruit) and two processed leaves during oolong tea post-harvest processing. We found that differential 5mC methylation among four key tissues is closely related to tissue functional differentiation and that genes expressed tissue-specifically, responsible for tissue-specific functions, maintain relatively low 5mC methylation levels relative to non-tissue-specifically expressed genes. Importantly, hypomethylation modifications of CsAlaDC and TS/GS genes in roots provided the molecular basis for the dominant synthesis of theanine in roots. In addition, integration of 5mC DNA methylationomics, metabolomics, and transcriptomics of post-harvest leaves revealed that content changes in flavor metabolites during oolong tea processing were closely associated with transcription level changes in corresponding metabolite synthesis genes, and changes in transcript levels of these important synthesis genes were strictly regulated by 5mC methylation. We further report that some key genes during processing are regulated by 5mC methylation, which can effectively explain the content changes of important aroma metabolites, including α-farnesene, nerolidol, lipids, and taste substances such as catechins. Our results not only highlight the key roles of 5mC methylation in important flavor substance synthesis in pre- and post-harvest processing, but also provide epimutation-related gene targets for future improvement of tea quality or breeding of whole-tissue high-theanine varieties.

Recent Advances in Assembly of Complex Plant Genomes.

Over the past 20 years, tremendous advances in sequencing technologies and computational algorithms have spurred plant genomic research into a thriving era with hundreds of genomes decoded already, ranging from those of nonvascular plants to those of flowering plants. However, complex plant genome assembly is still challenging and remains difficult to fully resolve with conventional sequencing and assembly methods due to high heterozygosity, highly repetitive sequences, or high ploidy characteristics of complex genomes. Herein, we summarize the challenges of and advances in complex plant genome assembly, including feasible experimental strategies, upgrades to sequencing technology, existing assembly methods, and different phasing algorithms. Moreover, we list actual cases of complex genome projects for readers to refer to and draw upon to solve future problems related to complex genomes. Finally, we expect that the accurate, gapless, telomere-to-telomere, and fully phased assembly of complex plant genomes could soon become routine.

Whole-genome Duplication Reshaped Adaptive Evolution in A Relict Plant Species, Cyclocarya paliurus.

Cyclocarya paliurus is a relict plant species that survived the last glacial period and shows a population expansion recently. Its leaves have been traditionally used to treat obesity and diabetes with the well-known active ingredient cyclocaric acid B. Here, we presented three C. paliurus genomes from two diploids with different flower morphs and one haplotype-resolved tetraploid assembly. Comparative genomic analysis revealed two rounds of recent whole-genome duplication events and identified 691 genes with dosage effects that likely contribute to adaptive evolution through enhanced photosynthesis and increased accumulation of triterpenoids. Resequencing analysis of 45 C. paliurus individuals uncovered two bottlenecks, consistent with the known events of environmental changes, and many selectively swept genes involved in critical biological functions, including plant defense and secondary metabolite biosynthesis. We also proposed the biosynthesis pathway of cyclocaric acid B based on multi-omics data and identified key genes, in particular gibberellin-related genes, associated with the heterodichogamy in C. paliurus species. Our study sheds light on evolutionary history of C. paliurus and provides genomic resources to study the medicinal herbs.

Pan-transcriptome assembly combined with multiple association analysis provides new insights into the regulatory network of specialized metabolites in the tea plant Camellia sinensis.

Specialized metabolites not only play important roles in biotic and abiotic stress adaptation of tea plants (Camellia sinensis (L.) O. Kuntze) but also contribute to the unique flavor of tea, the most important nonalcoholic beverage. However, the molecular networks and major genes that regulate specialized metabolites in tea plants are not well understood. Here, we constructed a population-level pan-transcriptome of the tea plant leaf using second-leaf transcriptome data from 134 accessions to investigate global expression differences in the population, expression presence or absence variations (ePAVs), and differentially expressed genes (DEGs) between pure Camellia sinensis var. assamica (CSA) and pure Camellia sinensis var. sinensis (CSS) accessions. Next, we used a genome-wide association study, a quantitative trait transcript study, and a transcriptome-wide association study to integrate genotypes, accumulation levels of specialized metabolites, and expression levels of pan-transcriptome genes to identify candidate regulatory genes for flavor-related metabolites and to construct a regulatory network for specialized metabolites in tea plants. The pan-transcriptome contains 30 482 expressed genes, 4940 and 5506 of which were newly annotated from a de novo transcriptome assembly without a reference and a genome reference-based assembly, respectively. DEGs and ePAVs indicated that CSA and CSS were clearly differentiated at the population transcriptome level, and they were closely related to abiotic tolerance and secondary metabolite synthesis phenotypes of CSA and CSS based on gene annotations. The regulatory network contained 212 specialized metabolites, 3843 candidate genes, and 3407 eQTLs, highlighting many pleiotropic candidate genes, candidate gene-rich eQTLs, and potential regulators of specialized metabolites. These included important transcription factors in the AP2/ERF-ERF, MYB, WD40, and bHLH families. CsTGY14G0001296, an ortholog of AtANS, appeared to be directly related to variation in proanthocyanins in the tea plant population, and the CsTGY11G0002074 gene encoding F3'5'H was found to contribute to the biased distribution of catechins between pure CSAs and pure CSSs. Together, these results provide a new understanding of the metabolite diversity in tea plants and offer new insights for more effective breeding of better-flavored tea varieties.

The spinach YY genome reveals sex chromosome evolution, domestication, and introgression history of the species.

BACKGROUND: Spinach (Spinacia oleracea L.) is a dioecious species with an XY sex chromosome system, but its Y chromosome has not been fully characterized. Our knowledge about the history of its domestication and improvement remains limited. RESULTS: A high-quality YY genome of spinach is assembled into 952 Mb in six pseudo-chromosomes. By a combination of genetic mapping, Genome-Wide Association Studies, and genomic analysis, we characterize a 17.42-Mb sex determination region (SDR) on chromosome 1. The sex chromosomes of spinach evolved when an insertion containing sex determination genes occurred, followed by a large genomic inversion about 1.98 Mya. A subsequent burst of SDR-specific repeats (0.1-0.15 Mya) explains the large size of this SDR. We identify a Y-specific gene, NRT1/PTR 6.4 which resides in this insertion, as a strong candidate for the sex determination or differentiation factor. Resequencing of 112 spinach genomes reveals a severe domestication bottleneck approximately 10.87 Kya, which dates the domestication of spinach 7000 years earlier than the archeological record. We demonstrate that a strong selection signal associated with internode elongation and leaf area expansion is associated with domestication of edibility traits in spinach. We find that several strong genomic introgressions from the wild species Spinacia turkestanica and Spinacia tetrandra harbor desirable alleles of genes related to downy mildew resistance, frost resistance, leaf morphology, and flowering-time shift, which likely contribute to spinach improvement. CONCLUSIONS: Analysis of the YY genome uncovers evolutionary forces shaping nascent sex chromosome evolution in spinach. Our findings provide novel insights about the domestication and improvement of spinach.

Signatures of selection in recently domesticated macadamia.

Macadamia is a high value nut crop that is recently domesticated, ideal for testing the effect of artificial selection. Here, we sequence the genome of Hawaiian cultivar 'Kau' and assemble into 794 Mb in 14 pseudo-chromosomes with 37,728 genes. Genome analysis reveals a whole-genome duplication event, occurred 46.8 million years ago. Gene expansions occurred in gene families involves in fatty acid biosynthesis. Gene duplication of MADS-Box transcription factors in proanthocyanidin biosynthesis are relevant for seed coat development. Genome re-sequencing of 112 accessions reveals the origin of Hawaiian cultivars from Mount Bauple in southeast Queensland in Australia. Selective sweeps are detected in macadamia cultivars, including genes involved in fatty acid biosynthesis, seed coat development, and heat stress response. Such strong effects of artificial selection in few generations reveals the genomic basis for 'one-step operation' for clonal crop domestication. The knowledge gained could accelerate domestication of new crops from wild species.

Uncovering the Novel QTLs and Candidate Genes of Salt Tolerance in Rice with Linkage Mapping, RTM-GWAS, and RNA-seq.

Salinity is a major abiotic stress that limits plant growth and crop productivity. Indica rice and japonica rice show significant differences in tolerance to abiotic stress, and it is considered a feasible method to breed progeny with stronger tolerance to abiotic stress by crossing indica and japonica rice. We herein developed a high-generation recombinant inbred lines (RILs) from Luohui 9 (indica) X RPY geng (japonica). Based on the high-density bin map of this RILs population, salt tolerance QTLs controlling final survival rates were analyzed by linkage mapping and RTM-GWAS methods. A total of seven QTLs were identified on chromosome 3, 4, 5, 6, and 8. qST-3.1, qST-5.1, qST-6.1, and qST-6.2 were novel salt tolerance QTLs in this study and their function were functionally verified by comparative analysis of parental genotype RILs. The gene aggregation result of these four new QTLs emphasized that the combination of the four QTL synergistic genotypes can significantly improve the salt stress tolerance of rice. By comparing the transcriptomes of the root tissues of the parents' seedlings, at 3 days and 7 days after salt treatment, we then achieved fine mapping of QTLs based on differentially expressed genes (DEGs) identification and DEGs annotations, namely, LOC_Os06g01250 in qST-6.1, LOC_Os06g37300 in qST-6.2, LOC_Os05g14880 in qST-5.1. The homologous genes of these candidate genes were involved in abiotic stress tolerance in different plants. These results indicated that LOC_Os05g14880, LOC_Os06g01250, and LOC_Os06g37300 were the candidate genes of qST-5.1, qST-6.1, and qST-6.2. Our finding provided novel salt tolerance-related QTLs, candidate genes, and several RILs with better tolerance, which will facilitate breeding for improved salt tolerance of rice varieties and promote the exploration tolerance mechanisms of rice salt stress.

Rambutan genome revealed gene networks for spine formation and aril development.

Rambutan is a popular tropical fruit known for its exotic appearance, has long flexible spines on shells, extraordinary aril growth, desirable nutrition, and a favorable taste. The genome of an elite rambutan cultivar Baoyan 7 was assembled into 328 Mb in 16 pseudo-chromosomes. Comparative genomics analysis between rambutan and lychee revealed that rambutan chromosomes 8 and 12 are collinear with lychee chromosome 1, which resulted in a chromosome fission event in rambutan (n = 16) or a fusion event in lychee (n = 15) after their divergence from a common ancestor 15.7 million years ago. Root development genes played a crucial role in spine development, such as endoplasmic reticulum pathway genes, jasmonic acid response genes, vascular bundle development genes, and K+ transport genes. Aril development was regulated by D-class genes (STK and SHP1), plant hormone and phenylpropanoid biosynthesis genes, and sugar metabolism genes. The lower rate of male sterility of hermaphroditic flowers appears to be regulated by MYB24. Population genomic analyses revealed genes in selective sweeps during domestication that are related to fruit morphology and environment stress response. These findings enhance our understanding of spine and aril development and provide genomic resources for rambutan improvement.

Haplotype-resolved genome assembly provides insights into evolutionary history of the tea plant Camellia sinensis.

Tea is an important global beverage crop and is largely clonally propagated. Despite previous studies on the species, its genetic and evolutionary history deserves further research. Here, we present a haplotype-resolved assembly of an Oolong tea cultivar, Tieguanyin. Analysis of allele-specific expression suggests a potential mechanism in response to mutation load during long-term clonal propagation. Population genomic analysis using 190 Camellia accessions uncovered independent evolutionary histories and parallel domestication in two widely cultivated varieties, var. sinensis and var. assamica. It also revealed extensive intra- and interspecific introgressions contributing to genetic diversity in modern cultivars. Strong signatures of selection were associated with biosynthetic and metabolic pathways that contribute to flavor characteristics as well as genes likely involved in the Green Revolution in the tea industry. Our results offer genetic and molecular insights into the evolutionary history of Camellia sinensis and provide genomic resources to further facilitate gene editing to enhance desirable traits in tea crops.

Structural variations in papaya genomes.

BACKGROUND: Structural variations (SVs) are a type of mutations that have not been widely detected in plant genomes and studies in animals have shown their role in the process of domestication. An in-depth study of SVs will help us to further understand the impact of SVs on the phenotype and environmental adaptability during papaya domestication and provide genomic resources for the development of molecular markers. RESULTS: We detected a total of 8083 SVs, including 5260 deletions, 552 tandem duplications and 2271 insertions with deletion being the predominant, indicating the universality of deletion in the evolution of papaya genome. The distribution of these SVs is non-random in each chromosome. A total of 1794 genes overlaps with SV, of which 1350 genes are expressed in at least one tissue. The weighted correlation network analysis (WGCNA) of these expressed genes reveals co-expression relationship between SVs-genes and different tissues, and functional enrichment analysis shows their role in biological growth and environmental responses. We also identified some domesticated SVs genes related to environmental adaptability, sexual reproduction, and important agronomic traits during the domestication of papaya. Analysis of artificially selected copy number variant genes (CNV-genes) also revealed genes associated with plant growth and environmental stress. CONCLUSIONS: SVs played an indispensable role in the process of papaya domestication, especially in the reproduction traits of hermaphrodite plants. The detection of genome-wide SVs and CNV-genes between cultivated gynodioecious populations and wild dioecious populations provides a reference for further understanding of the evolution process from male to hermaphrodite in papaya.

Chromosome-level reference genome assembly provides insights into aroma biosynthesis in passion fruit (Passiflora edulis).

Passion fruit, native to tropical America, is an agriculturally, economically and ornamentally important fruit plant that is well known for its acid pulp, rich aroma and distinctive flavour. Here, we present a chromosome-level genome assembly of passion fruit by incorporating PacBio long HiFi reads and Hi-C technology. The assembled reference genome is 1.28 Gb size with a scaffold N50 of 126.4 Mb and 99.22% sequences anchored onto nine pseudochromosomes. This genome is highly repetitive, accounting for 86.61% of the assembled genome. A total of 39,309 protein-coding genes were predicted with 93.48% of those being functionally annotated in the public databases. Genome evolution analysis revealed a core eudicot-common γ whole-genome triplication event and a more recent whole-genome duplication event, possibly contributing to the expansion of certain gene families. The 33 rapidly expanded gene families were significantly enriched in the pathways of isoflavone biosynthesis, galactose metabolism, diterpene biosynthesis and fatty acid metabolism, which might be responsible for the formation of featured flavours in the passion fruit. Transcriptome analysis revealed that genes related to ester and ethylene biosynthesis were significantly upregulated in the mature fruit and the expression levels of those genes were consistent with the accumulation of volatile lipid compounds. The passion fruit genome analysis improves our understanding of the genome evolution of this species and sheds new lights into the molecular mechanism of aroma biosynthesis in passion fruit.

Genomes of the Banyan Tree and Pollinator Wasp Provide Insights into Fig-Wasp Coevolution.

Banyan trees are distinguished by their extraordinary aerial roots. The Ficus genus includes species that have evolved a species-specific mutualism system with wasp pollinators. We sequenced genomes of the Chinese banyan tree, F. microcarpa, and a species lacking aerial roots, F. hispida, and one wasp genome coevolving with F. microcarpa, Eupristina verticillata. Comparative analysis of the two Ficus genomes revealed dynamic karyotype variation associated with adaptive evolution. Copy number expansion of auxin-related genes from duplications and elevated auxin production are associated with aerial root development in F. microcarpa. A male-specific AGAMOUS paralog, FhAG2, was identified as a candidate gene for sex determination in F. hispida. Population genomic analyses of Ficus species revealed genomic signatures of morphological and physiological coadaptation with their pollinators involving terpenoid- and benzenoid-derived compounds. These three genomes offer insights into and genomic resources for investigating the geneses of aerial roots, monoecy and dioecy, and codiversification in a symbiotic system.

Assembly of allele-aware, chromosomal-scale autopolyploid genomes based on Hi-C data.

Construction of chromosome-level assembly is a vital step in achieving the goal of a 'Platinum' genome, but it remains a major challenge to assemble and anchor sequences to chromosomes in autopolyploid or highly heterozygous genomes. High-throughput chromosome conformation capture (Hi-C) technology serves as a robust tool to dramatically advance chromosome scaffolding; however, existing approaches are mostly designed for diploid genomes and often with the aim of reconstructing a haploid representation, thereby having limited power to reconstruct chromosomes for autopolyploid genomes. We developed a novel algorithm (ALLHiC) that is capable of building allele-aware, chromosomal-scale assembly for autopolyploid genomes using Hi-C paired-end reads with innovative 'prune' and 'optimize' steps. Application on simulated data showed that ALLHiC can phase allelic contigs and substantially improve ordering and orientation when compared to other mainstream Hi-C assemblers. We applied ALLHiC on an autotetraploid and an autooctoploid sugar-cane genome and successfully constructed the phased chromosomal-level assemblies, revealing allelic variations present in these two genomes. The ALLHiC pipeline enables de novo chromosome-level assembly of autopolyploid genomes, separating each allele. Haplotype chromosome-level assembly of allopolyploid and heterozygous diploid genomes can be achieved using ALLHiC, overcoming obstacles in assembling complex genomes.