High-throughput Chromosome Conformation Capture Technology Research Articles

A chromosome-level genome assembly for Chinese plum ‘Wushancuili’ reveals the molecular basis of its fruit color and susceptibility to rain-cracking

Chinese plum (Prunus salicina Lindl.) originates from China and makes a large contribution to the global production of plums. The P. salicina ‘Wushancuili’ has a green coloration and high fruit quality and is economically important in eliminating poverty and protecting ecology in the Yangtze River Three Gorges Reservoir. However, rain-induced cracking (rain-cracking, literally skin cracking caused by rain) is a limitation to ‘Wushancuili’ fruit production and causes severe losses. This study reported a high-quality ‘Wushancuili’ genome assembly consisting of a 302.17-Mb sequence with eight pseudo-chromosomes and a contig N50 of 23.59 Mb through the combination of Illumina sequencing, Pacific Biosciences HiFi Ⅲ sequencing, and high-throughput chromosome conformation capture technology. A total of 25 109 protein-coding genes are predicted and 54.17% of the genome is composed of repetitive sequences. ‘Wushancuili’ underwent a remarkable orthoselection during evolution. Gene identification revealed that loss-of-function in four core MYB10 genes results in the anthocyanin deficiency and absence of red color, revealing the green coloration due to the residual high chlorophyll in fruit skin. Besides, the occurrence of cracking is assumed to be closely associated with cell wall modification and frequently rain-induced pathogen enrichment through transcriptomic analysis. The loss of MYB10 genes might render fruit more susceptible to pathogen-mediated cracking by weakening the epidermal strength and reactive oxygen species (ROS) scavenging. Our findings provided fundamental knowledge regarding fruit coloration and rain-cracking and will facilitate genetic improvement and cultivation management in Chinese plums.

Application of Hi-C technology in three-dimensional genomics research and disease pathogenesis analysis.

3D genomics aims to investigate the spatial structure of chromatin in the nucleus on the basis of genomic sequences, gene structures and relevant regulatory elements. The spatial organization of chromosomes is fundamental for gene expression regulation. Recent advances of high-throughput chromosome conformation capture (Hi-C) technology and its derivatives, has enabled capture of chromatin architecture with high resolution. In this review, we summarize the development and applications of various technologies of 3D genomes in disease research, particularly in the elucidation of pathogenic mechanisms in cancers and other systemic disorders.

High-quality haplotype-resolved genome assembly of cultivated octoploid strawberry.

Cultivated strawberry (Fragaria × ananassa), a perennial herb belonging to the family Rosaceae, is a complex octoploid with high heterozygosity at most loci. However, there is no research on the haplotype of the octoploid strawberry genome. Here we aimed to obtain a high-quality genome of the cultivated strawberry cultivar, "Yanli", using single molecule real-time sequencing and high-throughput chromosome conformation capture technology. The "Yanli" genome was 823Mb in size, with a long terminal repeat assembly index of 14.99. The genome was phased into two haplotypes, Hap1 (825Mb with contig N50 of 26.70Mb) and Hap2 (808Mb with contig N50 of 27.51Mb). Using the combination of Hap1 and Hap2, we obtained for the first time a haplotype-resolved genome with 56 chromosomes for the cultivated octoploid strawberry. We identified a ~ 10Mb inversion and translocation on chromosome 2-1. 104 957 and 102 356 protein-coding genes were annotated in Hap1 and Hap2, respectively. Analysis of the genes related to the anthocyanin biosynthesis pathway revealed the structural diversity and complexity in the expression of the alleles in the octoploid F. × ananassa genome. In summary, we obtained a high-quality haplotype-resolved genome assembly of F. × ananassa, which will provide the foundation for investigating gene function and evolution of the genome of cultivated octoploid strawberry.

Genome Resource for the Verticillium Wilt Resistant Gossypium hirsutum Cultivar Zhongzhimian No. 2.

Verticillium wilt, caused by the fungal pathogen Verticillium dahliae, is the major cause of disease-related yield losses in cotton (Gossypium hirsutum). Despite these losses, the major cultivars of G. hirsutum remain highly susceptible to Verticillium wilt. The lack of understanding on the genetic basis for Verticillium wilt resistance may further hinder progress in deploying elite cultivars with proven resistance, such as the wilt resistant G. hirsutum cultivar Zhongzhimian No. 2. To help remedy this knowledge gap, we sequenced the whole genome of Zhongzhimian No. 2 and assembled it from a combination of PacBio long reads, Illumina short reads, and high-throughput chromosome conformation capture technologies. The final assembly of the genome was 2.33 Gb, encoding 95,327 predicted coding sequences. The GC content was 34.39% with 99.2% of the bases anchored to 26 pseudo-chromosomes that ranged from 53.8 to 127.7 Mb. This resource will help gain a detailed understanding of the genomic features governing high yield and Verticillium wilt resistance in this cultivar. Comparative genomics will be particularly helpful, since there are several published genomes of other Gossypium species. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Chromosome-scale genome assembly of Camellia sinensis combined with multi-omics provides insights into its responses to infestation with green leafhoppers.

The tea plant (Camellia sinensis) is an important economic crop, which is becoming increasingly popular worldwide, and is now planted in more than 50 countries. Tea green leafhopper is one of the major pests in tea plantations, which can significantly reduce the yield and quality of tea during the growth of plant. In this study, we report a genome assembly for DuyunMaojian tea plants using a combination of Oxford Nanopore Technology PromethION™ with high-throughput chromosome conformation capture technology and used multi-omics to study how the tea plant responds to infestation with tea green leafhoppers. The final genome was 3.08 Gb. A total of 2.97 Gb of the genome was mapped to 15 pseudo-chromosomes, and 2.79 Gb of them could confirm the order and direction. The contig N50, scaffold N50 and GC content were 723.7 kb, 207.72 Mb and 38.54%, respectively. There were 2.67 Gb (86.77%) repetitive sequences, 34,896 protein-coding genes, 104 miRNAs, 261 rRNA, 669 tRNA, and 6,502 pseudogenes. A comparative genomics analysis showed that DuyunMaojian was the most closely related to Shuchazao and Yunkang 10, followed by DASZ and tea-oil tree. The multi-omics results indicated that phenylpropanoid biosynthesis, α-linolenic acid metabolism, flavonoid biosynthesis and 50 differentially expressed genes, particularly peroxidase, played important roles in response to infestation with tea green leafhoppers (Empoasca vitis Göthe). This study on the tea tree is highly significant for its role in illustrating the evolution of its genome and discovering how the tea plant responds to infestation with tea green leafhoppers will contribute to a theoretical foundation to breed tea plants resistant to insects that will ultimately result in an increase in the yield and quality of tea.

The genetic adaptations of Toxoptera aurantii facilitated its rapid multiple plant hosts dispersal and invasion

Toxoptera aurantii Boyer de Fonscolombe (Hemiptera: Aphididae) can attack many plant hosts, including tea (Camellia sinensis L.), citrus (Citrus spp.), lychee (Litchi chinensis Sonn.), banana (Musa spp.), and pineapple (Ananas comasus L.) among others. It is a widely distributed hexapod and one of the most destructive pests in tea plantations, causing enormous economic losses in tea production each year. A high-quality reference genome is important to study the phylogenetics and evolution of T. aurantii because its genome is highly heterozygous and repetitive. We obtained a de novo genome assembly of T. aurantii at the chromosome level using a combination of long Nanopore reads from sequencing with high-throughput chromosome conformation capture technology. When finally assembled, the genome was 318.95 Mb on four chromosomes with a 15.19 Mb scaffold N50. A total of 12,162 genes encoded proteins, while there were 22.01% repetitive sequences that totaled 67.73 Mb. Phylogenetic analyses revealed that T. aurantii and Aphis gossypii parted ways approximately 7.6 million years ago (Mya). We used a combination of long-read single-molecule sequencing with Hi-C–based chromatin interaction maps that resulted in a reference chromosomal level reference genome of T. aurantii that was high quality. Our results will enable the exploration of the genetics behind the special biological features of T. aurantii and also provide a source of data that should be useful to compare the compare genome among the Hemiptera.

Chromosome‐level genome of Entada phaseoloides provides insights into genome evolution and biosynthesis of triterpenoid saponins

As a medicinal herbal plant, Entada phaseoloides has high levels of secondary metabolites, particularly triterpenoid saponins, which are important resources for scientific research and medical applications. However, the lack of a reference genome for this genus has limited research on its evolution and utilization of its medicinal potential. In this study, we report a chromosome-scale genome assembly for E. phaseoloides using Illumina, Nanopore long reads and high-throughput chromosome conformation capture technology. The assembled reference genome is 456.18 Mb (scaffold N50=30.9Mb; contig N50=6.34 Mb) with 95.71% of the sequences anchored onto 14 pseudochromosomes. E. phaseoloides was estimated to have diverged from the Leguminosae lineage at ~72.0 million years ago. With the integration of transcriptomic and metabolomic data, gene expression patterns and metabolite profiling of E. phaseoloides were determined in different tissues. The pattern of gene expression and metabolic profile of the kernel were distinct from those of other tissues. Furthermore, the evolution of certain gene families involved in the biosynthesis of triterpenoid saponins and terpenes was analysed and offers new insights into the formation of these two metabolites. Four CYP genes, one UGT gene and related transcription factors were identified as candidate genes contributing to regulation of triterpenoid saponin biosynthesis. As the first high-quality assembled reference genome in the genus Entada, it will not only provide new information for the evolutionary study of this genus and conservation biology of E. phaseoloides but also lay a foundation for the formation and utilization of secondary metabolites in medicinal plants.

Chromosomal-Level Assembly of Antarctic Scaly Rockcod, Trematomus loennbergii Genome Using Long-Read Sequencing and Chromosome Conformation Capture (Hi-C) Technologies

Trematomus species (suborder Notothenioidei; family Nototheniidae) are widely distributed in the southern oceans near Antarctica. There are 11 recognized species in the genus Trematomus, and notothenioids are known to have high chromosomal diversity (2n = 24–58) because of relatively recent and rapid adaptive radiation. Herein, we report the chromosomal-level genome assembly of T. loennbergii, the first characterized genome representative of the genus Trematomus. The final genome assembly of T. loennbergii was obtained using a Pacific Biosciences long-read sequencing platform and high-throughput chromosome conformation capture technology. Twenty-three chromosomal-level scaffolds were assembled to 940 Mb in total size, with a longest contig size of 48.5 Mb and contig N50 length of 24.7 Mb. The genome contained 42.03% repeat sequences, and a total of 24,525 protein-coding genes were annotated. We produced a high-quality genome assembly of T. loennbergii. Our results provide a first reference genome for the genus Trematomus and will serve as a basis for studying the molecular taxonomy and evolution of Antarctic fish.

Chromosome-level genome assembly of the hard-shelled mussel Mytilus coruscus, a widely distributed species from the temperate areas of East Asia.

BackgroundThe hard-shelled mussel (Mytilus coruscus) is widely distributed in the temperate seas of East Asia and is an important commercial bivalve in China. Chromosome-level genome information of this species will contribute not only to the development of hard-shelled mussel genetic breeding but also to studies on larval ecology, climate change biology, marine biology, aquaculture, biofouling, and antifouling.FindingsWe applied a combination of Illumina sequencing, Oxford Nanopore Technologies sequencing, and high-throughput chromosome conformation capture technologies to construct a chromosome-level genome of the hard-shelled mussel, with a total length of 1.57 Gb and a median contig length of 1.49 Mb. Approximately 90.9% of the assemblies were anchored to 14 linkage groups. We assayed the genome completeness using BUSCO. In the metazoan dataset, the present assemblies have 89.4% complete, 1.9% incomplete, and 8.7% missing BUSCOs. Gene modeling enabled the annotation of 37,478 protein-coding genes and 26,917 non-coding RNA loci. Phylogenetic analysis showed that M. coruscus is the sister taxon to the clade including Modiolus philippinarum and Bathymodiolus platifrons. Conserved chromosome synteny was observed between hard-shelled mussel and king scallop, suggesting that this is shared ancestrally. Transcriptomic profiling indicated that the pathways of catecholamine biosynthesis and adrenergic signaling in cardiomyocytes might be involved in metamorphosis.ConclusionsThe chromosome-level assembly of the hard-shelled mussel genome will provide novel insights into mussel genome evolution and serve as a fundamental platform for studies regarding the planktonic-sessile transition, genetic diversity, and genomic breeding of this bivalve.

Exploring chromatin conformation and gene co-expression through graph embedding.

The relationship between gene co-expression and chromatin conformation is of great biological interest. Thanks to high-throughput chromosome conformation capture technologies (Hi-C), researchers are gaining insights on the tri-dimensional organization of the genome. Given the high complexity of Hi-C data and the difficult definition of gene co-expression networks, the development of proper computational tools to investigate such relationship is rapidly gaining the interest of researchers. One of the most fascinating questions in this context is how chromatin topology correlates with gene co-expression and which physical interaction patterns are most predictive of co-expression relationships. To address these questions, we developed a computational framework for the prediction of co-expression networks from chromatin conformation data. We first define a gene chromatin interaction network where each gene is associated to its physical interaction profile; then, we apply two graph embedding techniques to extract a low-dimensional vector representation of each gene from the interaction network; finally, we train a classifier on gene embedding pairs to predict if they are co-expressed. Both graph embedding techniques outperform previous methods based on manually designed topological features, highlighting the need for more advanced strategies to encode chromatin information. We also establish that the most recent technique, based on random walks, is superior. Overall, our results demonstrate that chromatin conformation and gene regulation share a non-linear relationship and that gene topological embeddings encode relevant information, which could be used also for downstream analysis. The source code for the analysis is available at: https://github.com/marcovarrone/gene-expression-chromatin. Supplementary data are available at Bioinformatics online.

High-quality genome assembly of Metaphire vulgaris.

Earthworms enrich the soil and protect the health of their ecological environment. Previous studies on these invertebrates determined their protein content, hormone secretions, medicinal value, and ecological habits, but their whole genomic sequence remains incomplete. We performed whole genome sequencing of Metaphire vulgaris (Chen, 1930), which belongs to the genus Metaphire of the family Megascolecidae. The genome assembly was 729 Mb, with a N50 contig size of 4.2 Mb. In total, 559 contigs were anchored to 41 chromosomes according to the results of Hi-C (High-throughput Chromosome Conformation Capture) technology, which was confirmed by karyological analysis. A comparison of the genomic sequences and genes indicated that there was a whole-genome duplication in M. vulgaris followed by several chromosome fusion events. Hox genes and lumbrokinase genes were identified as partial clusters surrounding the genome. Our high-quality genome assembly of M. vulgaris will provide valuable information for gene function and evolutionary studies in earthworms.

Chromosome-Level Clam Genome Helps Elucidate the Molecular Basis of Adaptation to a Buried Lifestyle.

SummaryBivalve mollusks are economically important invertebrates that exhibit marked diversity in benthic lifestyle and provide valuable resources for understanding the molecular basis of adaptation to benthic life. In this report, we present a high-quality, chromosome-anchored reference genome of the Venus clam, Cyclina sinensis. The chromosome-level genome was assembled by Pacific Bioscience single-molecule real-time sequencing, Illumina paired-end sequencing, 10× Genomics, and high-throughput chromosome conformation capture technologies. The final genome assembly of C. sinensis is 903.2 Mb in size, with a contig N50 size of 2.6 Mb and a scaffold N50 size of 46.5 Mb. Enrichment analyses of significantly expanded and positively selected genes suggested evolutionary adaptation of this clam to buried life. In addition, a change in shell color represents another mechanism of adaptation to burial in sediment. The high-quality genome generated in this work provides a valuable resource for investigating the molecular mechanisms of adaptation to buried lifestyle.

Gossypium Genomics: Trends, Scope, and Utilization for Cotton Improvement.

Cotton (Gossypium spp.) is the most important natural fiber crop worldwide. The diversity of Gossypium species also provides an ideal model for investigating evolution and domestication of polyploids. However, the huge and complex cotton genome hinders genomic research. Technical advances in high-throughput sequencing and bioinformatics analysis have now largely overcome these obstacles, bringing about a new era of cotton genomics. Here, we review recent progress in Gossypium genomics based on whole genome sequencing, resequencing, and comparative genomics, which have provided insights about the genomic basis of fiber biogenesis and the landscape of cotton functional genomics. We address current challenges and present multidisciplinary genomics-enabled breeding strategies covering the breadth of high fiber yield, quality, and environmental resilience for future cotton breeding programs.

Chromosomal-level reference genome of Chinese peacock butterfly (Papilio bianor) based on third-generation DNA sequencing and Hi-C analysis.

Background Papilio bianor Cramer, 1777 (commonly known as the Chinese peacock butterfly) (Insecta, Lepidoptera, Papilionidae) is a widely distributed swallowtail butterfly with a wide number of geographic populations ranging from the southeast of Russia to China, Japan, India, Vietnam, Myanmar, and Thailand. Its wing color consists of both pigmentary colored scales (black, reddish) and structural colored scales (iridescent blue or green dust). A high-quality reference genome of P. bianor is an important foundation for investigating iridescent color evolution, phylogeography, and the evolution of swallowtail butterflies.FindingsWe obtained a chromosome-level de novo genome assembly of the highly heterozygous P. bianor using long Pacific Biosciences sequencing reads and high-throughput chromosome conformation capture technology. The final assembly is 421.52 Mb on 30 chromosomes (29 autosomes and 1 Z sex chromosome) with 13.12 Mb scaffold N50. In total, 15,375 protein-coding genes and 233.09 Mb of repetitive sequences were identified. Phylogenetic analyses indicated that P. bianor separated from a common ancestor of swallowtails ∼23.69–36.04 million years ago. Demographic history suggested that the population expansion of this species from the last interglacial period to the last glacial maximum possibly resulted from its decreased natural enemies and its adaptation to climate change during the glacial period.ConclusionsWe present a high-quality chromosome-level reference genome of P. bianor using long-read single-molecule sequencing and Hi-C–based chromatin interaction maps. Our results lay the foundation for exploring the genetic basis of special biological features of P. bianor and also provide a useful data source for comparative genomics and phylogenomics among butterflies and moths.

Chromosome-level assembly of the mustache toad genome using third-generation DNA sequencing and Hi-C analysis.

BackgroundThe mustache toad, Vibrissaphora ailaonica, is endemic to China and belongs to the Megophryidae family. Like other mustache toad species, V. ailaonica males temporarily develop keratinized nuptial spines on their upper jaw during each breeding season, which fall off at the end of the breeding season. This feature is likely result of the reversal of sexual dimorphism in body size, with males being larger than females. A high-quality reference genome for the mustache toad would be invaluable to investigate the genetic mechanism underlying these repeatedly developing keratinized spines.FindingsTo construct the mustache toad genome, we generated 225 Gb of short reads and 277 Gb of long reads using Illumina and Pacific Biosciences (PacBio) sequencing technologies, respectively. Sequencing data were assembled into a 3.53-Gb genome assembly, with a contig N50 length of 821 kb. We also used high-throughput chromosome conformation capture (Hi-C) technology to identify contacts between contigs, then assembled contigs into scaffolds and assembled a genome with 13 chromosomes and a scaffold N50 length of 412.42 Mb. Based on the 26,227 protein-coding genes annotated in the genome, we analyzed phylogenetic relationships between the mustache toad and other chordate species. The mustache toad has a relatively higher evolutionary rate and separated from a common ancestor of the marine toad, bullfrog, and Tibetan frog 206.1 million years ago. Furthermore, we identified 201 expanded gene families in the mustache toad, which were mainly enriched in immune pathway, keratin filament, and metabolic processes.ConclusionsUsing Illumina, PacBio, and Hi-C technologies, we constructed the first high-quality chromosome-level mustache toad genome. This work not only offers a valuable reference genome for functional studies of mustache toad traits but also provides important chromosomal information for wider genome comparisons.

Advances in mammalian three-dimensional genome by using Hi-C technology approach

Mammalian genomic DNA in the cell nucleus doesn't exist in linear form but is highly folded and condensed into chromatin with a three-dimensional (3D) structure possessing a specific spatial structure and conformation. Hi-C, the high-throughput chromosome conformation capture technology, was first published in 2009, and it provides an in-depth view of 3D genomics. According to the size of DNA unit, the 3D hierarchical units of mammalian genome can be categorized sequentially as chromosome territory (CT), chromatin compartment A/B, topological associated domain (TAD), and chromatin loop. These hierarchical structural units play vital roles in gene transcription and regulation. In this review, we summarize the 3D hierarchical division of chromosomes, the effects of hierarchical units and the applications of Hi-C technology in development and disease. This review is intended to provide insights for the further study of 3D genomics in mammals.

Sub-nucleosomal Genome Structure Reveals Distinct Nucleosome Folding Motifs

Elucidating the global and local rules that govern genome-wide, hierarchical chromatin architecture remains a critical challenge. Current high-throughput chromosome conformation capture (Hi-C) technologies have identified large-scale chromatin structural motifs, such as topologically associating domains and looping. However, structural rules at the smallest or nucleosome scale remain poorly understood. Here, we coupled nucleosome-resolved Hi-C technology with simulated annealing-molecular dynamics (SA-MD) simulation to reveal 3D spatial distributions of nucleosomes and their genome-wide orientation in chromatin. Our method, called Hi-CO, revealed distinct nucleosome folding motifs across the yeast genome. Our results uncovered two types of basic secondary structural motifs in nucleosome folding: α-tetrahedron and β-rhombus analogous to α helix and β sheet motifs in protein folding. Using mutants and cell-cycle-synchronized cells, we further uncovered motifs with specific nucleosome positioning and orientation coupled to epigenetic features at individual loci. By illuminating molecular-level structure-function relationships in eukaryotic chromatin, our findings establish organizational principles of nucleosome folding.

Genome of an allotetraploid wild peanut Arachis monticola: a de novo assembly.

Arachis monticola (2n = 4x = 40) is the only allotetraploid wild peanut within the Arachis genus and section, with an AABB-type genome of ∼2.7 Gb in size. The AA-type subgenome is derived from diploid wild peanut Arachis duranensis, and the BB-type subgenome is derived from diploid wild peanut Arachis ipaensis. A. monticola is regarded either as the direct progenitor of the cultivated peanut or as an introgressive derivative between the cultivated peanut and wild species. The large polyploidy genome structure and enormous nearly identical regions of the genome make the assembly of chromosomal pseudomolecules very challenging. Here we report the first reference quality assembly of the A. monticola genome, using a series of advanced technologies. The final whole genome of A. monticola is ∼2.62 Gb and has a contig N50 and scaffold N50 of 106.66 Kb and 124.92 Mb, respectively. The vast majority (91.83%) of the assembled sequence was anchored onto the 20 pseudo-chromosomes, and 96.07% of assemblies were accurately separated into AA- and BB- subgenomes. We demonstrated efficiency of the current state of the strategy for de novo assembly of the highly complex allotetraploid species, wild peanut (A. monticola), based on whole-genome shotgun sequencing, single molecule real-time sequencing, high-throughput chromosome conformation capture technology, and BioNano optical genome maps. These combined technologies produced reference-quality genome of the allotetraploid wild peanut, which is valuable for understanding the peanut domestication and evolution within the Arachis genus and among legume crops.

Multi-scale structural community organisation of the human genome

BackgroundStructural interaction frequency matrices between all genome loci are now experimentally achievable thanks to high-throughput chromosome conformation capture technologies. This ensues a new methodological challenge for computational biology which consists in objectively extracting from these data the structural motifs characteristic of genome organisation.ResultsWe deployed the fast multi-scale community mining algorithm based on spectral graph wavelets to characterise the networks of intra-chromosomal interactions in human cell lines. We observed that there exist structural domains of all sizes up to chromosome length and demonstrated that the set of structural communities forms a hierarchy of chromosome segments. Hence, at all scales, chromosome folding predominantly involves interactions between neighbouring sites rather than the formation of links between distant loci.ConclusionsMulti-scale structural decomposition of human chromosomes provides an original framework to question structural organisation and its relationship to functional regulation across the scales. By construction the proposed methodology is independent of the precise assembly of the reference genome and is thus directly applicable to genomes whose assembly is not fully determined.