Spatial proximity and gene function: a new dimension in prokaryotic gene association network analysis with 3D-GeneNet.

Understanding the biological functions and processes of genes, particularly those not yet characterized, is crucial for advancing molecular biology and identifying therapeutic targets. The hypothesis guiding this study is that the 3D proximity of genes correlates with their functional interactions and relevance in prokaryotes. We introduced 3D-GeneNet, an innovative software tool that utilizes high-throughput sequencing data from chromosome conformation capture techniques and integrates topological metrics to construct gene association networks. Through a series of comparative analyses focused on spatial versus linear distances, we explored various dimensions such as topological structure, functional enrichment levels, distribution patterns of linear distances among gene pairs, and the area under the receiver operating characteristic curve by utilizing model organism Escherichia coli K-12. Furthermore, 3D-GeneNet was shown to maintain good accuracy compared to multiple algorithms (neighbourhood, co-occurrence, coexpression, and fusion) across multiple bacteria, including E. coli, Brucella abortus, and Vibrio cholerae. In addition, the accuracy of 3D-GeneNet's prediction of long-distance gene interactions was identified by bacterial two-hybrid assays on E. coli K-12 MG1655, where 3D-GeneNet not only increased the accuracy of linear genomic distance tripled but also achieved 60% accuracy by running alone. Finally, it can be concluded that the applicability of 3D-GeneNet will extend to various bacterial forms, including Gram-negative, Gram-positive, single-, and multi-chromosomal bacteria through Hi-C sequencing and analysis. Such findings highlight the broad applicability and significant promise of this method in the realm of gene association network. 3D-GeneNet is freely accessible at https://github.com/gaoyuanccc/3D-GeneNet.

Distributions, interactions, and dynamics of prokaryotes and phages in a hybrid biological wastewater treatment system.

BACKGROUND: Understanding the interactions and dynamics of microbiotas within biological wastewater treatment systems is essential for ensuring their stability and long-term sustainability. In this study, we developed a systematic framework employing multi-omics and Hi-C sequencing to extensively investigate prokaryotic and phage communities within a hybrid biofilm and activated sludge system. RESULTS: We uncovered distinct distribution patterns, metabolic capabilities, and activities of functional prokaryotes through the analysis of 454 reconstructed prokaryotic genomes. Additionally, we reconstructed a phage catalog comprising 18,645 viral operational taxonomic units (vOTUs) with high length and contiguity using hybrid assembly, and a distinct distribution of phages was depicted between activated sludge (AS) and biofilm. Importantly, 1340 host-phage pairs were established using Hi-C and conventional in silico methods, unveiling the host-determined phage prevalence. The majority of predicted hosts were found to be involved in various crucial metabolic processes, highlighting the potential vital roles of phages in influencing substance metabolism within this system. Moreover, auxiliary metabolic genes (AMGs) related to various categories (e.g., carbohydrate degradation, sulfur metabolism, transporter) were predicted. Subsequent activity analysis emphasized their potential ability to mediate host metabolism during infection. We also profiled the temporal dynamics of phages and their associated hosts using 13-month time-series metagenomic data, further demonstrating their tight interactions. Notably, we observed lineage-specific infection patterns, such as potentially host abundance- or phage/host ratio-driven phage population changes. CONCLUSIONS: The insights gained from this research contribute to the growing body of knowledge surrounding interactions and dynamics of host-phage and pave the way for further exploration and potential applications in the field of microbial ecology. Video Abstract.

Exploring the cobia (Rachycentron canadum) genome: unveiling putative male heterogametic regions and identification of sex-specific markers.

BACKGROUND: Cobia (Rachycentron canadum) is the only member of the Rachycentridae family and exhibits considerable sexual dimorphism in growth rate. Sex determination in teleosts has been a long-standing basic biological question, and the molecular mechanisms of sex determination/differentiation in cobia are completely unknown. RESULTS: Here, we reported 2 high-quality, chromosome-level annotated male and female cobia genomes with assembly sizes of 586.51 Mb (contig/scaffold N50: 86.0 kb/24.3 Mb) and 583.88 Mb (79.9 kb/22.5 Mb), respectively. Synteny inference among perciform genomes revealed that cobia and the remora Echeneis naucrates were sister groups. Further, whole-genome resequencing of 31 males and 60 females, genome-wide association study, and sequencing depth analysis identified 3 short male-specific regions within a 10.7-kb continuous genomic region on male chromosome 18, which hinted at an undifferentiated sex chromosome system with a putative XX/XY mode of sex determination in cobia. Importantly, the only 2 genes within/between the male-specific regions, epoxide hydrolase 1 (ephx1, renamed cephx1y) and transcription factor 24 (tcf24, renamed ctcf24y), showed testis-specific/biased gene expression, whereas their counterparts cephx1x and ctf24x, located in female chromosome 18, were similarly expressed in both sexes. In addition, male-specific PCR targeting the cephx1y gene revealed that this genomic feature is conserved in cobia populations from Panama, Brazil, Australia, and Japan. CONCLUSION: The first comprehensive genomic survey presented here is a valuable resource for future studies on cobia population structure and dynamics, conservation, and evolutionary history. Furthermore, it establishes evidence of putative male heterogametic regions with 2 genes playing a potential role in the sex determination of the species, and it provides further support for the rapid evolution of sex-determining mechanisms in teleost fish.

Three-dimensional genome architecture persists in a 52,000-year-old woolly mammoth skin sample.

Analyses of ancient DNA typically involve sequencing the surviving short oligonucleotides and aligning to genome assemblies from related, modern species. Here, we report that skin from a female woolly mammoth (†Mammuthus primigenius) that died 52,000 years ago retained its ancient genome architecture. We use PaleoHi-C to map chromatin contacts and assemble its genome, yielding 28 chromosome-length scaffolds. Chromosome territories, compartments, loops, Barr bodies, and inactive X chromosome (Xi) superdomains persist. The active and inactive genome compartments in mammoth skin more closely resemble Asian elephant skin than other elephant tissues. Our analyses uncover new biology. Differences in compartmentalization reveal genes whose transcription was potentially altered in mammoths vs. elephants. Mammoth Xi has a tetradic architecture, not bipartite like human and mouse. We hypothesize that, shortly after this mammoth's death, the sample spontaneously freeze-dried in the Siberian cold, leading to a glass transition that preserved subfossils of ancient chromosomes at nanometer scale.

A chromosome-level reference genome assembly and a full-length transcriptome assembly of the giant freshwater prawn (Macrobrachium rosenbergii).

The giant freshwater prawn (Macrobrachium rosenbergii) is a key species in the aquaculture industry in several Asian, African and South American countries. Despite a considerable growth in its production worldwide, the genetic complexities of M. rosenbergii various morphotypes pose challenges in cultivation. This study reports the first chromosome-scale reference genome and a high-quality full-length transcriptome assembly for M. rosenbergii. We employed the PacBio High Fidelity (HiFi) sequencing to obtain an initial draft assembly and further scaffolded it with the chromatin contact mapping (Hi-C) technique to achieve a final assembly of 3.73-Gb with an N50 scaffold length of 33.6 Mb. Repetitive elements constituted nearly 60% of the genome assembly, with simple sequence repeats and retrotransposons being the most abundant. The availability of both the chromosome-scale assembly and the full-length transcriptome assembly enabled us to thoroughly probe alternative splicing events in M. rosenbergii. Among the 2,041 events investigated, exon skipping represented the most prevalent class, followed by intron retention. Interestingly, specific isoforms were observed across multiple tissues. Additionally, within a single tissue type, transcripts could undergo alternative splicing, yielding multiple isoforms. We believe that the availability of a chromosome-level reference genome for M. rosenbergii along with its full-length transcriptome will be instrumental in advancing our understanding of the giant freshwater prawn biology and enhancing its molecular breeding programs, paving the way for the development of M. rosenbergii with valuable traits in commercial aquaculture.

Shedding Light on Bacterial Chromosome Structure: Exploring the Significance of 3C-Based Approaches.

The complex architecture of DNA within living organisms is essential for maintaining the genetic information that dictates their functions and characteristics. Among the many complexities of genetic material organization, the folding and arrangement of DNA into chromosomes play a critical role in regulating gene expression, replication, and other essential cellular processes. Bacteria, despite their apparently simple cellular structure, exhibit a remarkable level of chromosomal organization that influences their adaptability and survival in diverse environments. Understanding the three-dimensional arrangement of bacterial chromosomes has long been a challenge due to technical limitations, but the development of Chromosome Conformation Capture (3C) methods revolutionized our ability to explore the hierarchical structure and the dynamics of bacterial genomes. Here, we review the major advances in the field of bacterial chromosome structure using 3C technology over the past decade.

High-Throughput Mapping of Chromosomal Conformations in E. coli Under Physiological Conditions Using Massively Multiplexed Mu Transposition.

Many approaches for measuring three-dimensional chromosomal conformations rely upon formaldehyde crosslinking followed by subsequent proximity ligation, a family of methods exemplified by 3C, Hi-C, etc. Here we provide an alternative crosslinking-free procedure for high-throughput identification of long-range contacts in the chromosomes of enterobacteria, making use of contact-dependent transposition of phage Mu to identify distant loci in close contact. The procedure described here will suffice to provide a comprehensive map of transposition frequencies between tens of thousands of loci in a bacterial genome, with the resolution limited by the diversity of the insertion site library used and the sequencing depth applied.

Motorized chain models of the ideal chromosome.

An array of motor proteins consumes chemical energy in setting up the architectures of chromosomes. Here, we explore how the structure of ideal polymer chains is influenced by two classes of motors. The first class which we call "swimming motors" acts to propel the chromatin fiber through three-dimensional space. They represent a caricature of motors such as RNA polymerases. Previously, they have often been described by adding a persistent flow onto Brownian diffusion of the chain. The second class of motors, which we call "grappling motors" caricatures the loop extrusion processes in which segments of chromatin fibers some distance apart are brought together. We analyze these models using a self-consistent variational phonon approximation to a many-body Master equation incorporating motor activities. We show that whether the swimming motors lead to contraction or expansion depends on the susceptibility of the motors, that is, how their activity depends on the forces they must exert. Grappling motors in contrast to swimming motors lead to long-ranged correlations that resemble those first suggested for fractal globules and that are consistent with the effective interactions inferred by energy landscape analyses of Hi-C data on the interphase chromosome.

Multiscale Bayesian simulations reveal functional chromatin condensation of gene loci.

Chromatin, the complex assembly of DNA and associated proteins, plays a pivotal role in orchestrating various genomic functions. To aid our understanding of the principles underlying chromatin organization, we introduce Hi-C metainference, a Bayesian approach that integrates Hi-C contact frequencies into multiscale prior models of chromatin. This approach combines both bottom-up (the physics-based prior) and top-down (the data-driven posterior) strategies to characterize the 3D organization of a target genomic locus. We first demonstrate the capability of this method to accurately reconstruct the structural ensemble and the dynamics of a system from contact information. We then apply the approach to investigate the Sox2, Pou5f1, and Nanog loci of mouse embryonic stem cells using a bottom-up chromatin model at 1 kb resolution. We observe that the studied loci are conformationally heterogeneous and organized as crumpled globules, favoring contacts between distant enhancers and promoters. Using nucleosome-resolution simulations, we then reveal how the Nanog gene is functionally organized across the multiple scales of chromatin. At the local level, we identify diverse tetranucleosome folding motifs with a characteristic distribution along the genome, predominantly open at cis-regulatory elements and compact in between. At the larger scale, we find that enhancer-promoter contacts are driven by the transient condensation of chromatin into compact domains stabilized by extensive internucleosome interactions. Overall, this work highlights the condensed, but dynamic nature of chromatin in vivo, contributing to a deeper understanding of gene structure-function relationships.

A chromosome-level genome assembly and annotation of the medicinal plant Lepidium apetalum.

OBJECTIVES: As a traditional Chinese medicine, Lepidium apetalum is commonly used for purging the lung, relieving dyspnea, alleviating edema, and has the significant pharmacological effects on cardiovascular disease, hyperlipidemia, etc. In addition, the seeds of L. apetalum are rich in unsaturated fatty acids, sterols, glucosinolates and have a variety of biological activity compounds. To facilitate genomics, phylogenetic and secondary metabolite biosynthesis studies of L. apetalum, we assembled the high-resolution genome of L. apetalum. DATA DESCRIPTION: We completed chromosome-level genome assembly of the L. apetalum genome (2n = 32), using Illumina HiSeq and PacBio Sequel sequencing platform as well as high-throughput chromosome conformation capture (Hi-C) technique. The assembled genome was 296.80 Mb in size, 34.41% in GC content, and 23.89% in repeated sequence content, including 316 contigs with a contig N50 of 16.31 Mb. Hi-C scaffolding resulted in 16 chromosomes occupying 99.79% of the assembled genome sequences. A total of 46 584 genes and 105 pseudogenes were predicted, 98.37% of which can be annotated to Nr, GO, KEGG, TrEMBL, SwissPort, Pfam and KOG databases. The high-quality reference genome generated by this study will provide accurate genetic information for the molecular biology research of L. apetalum.

C2c: Predicting Micro-C from Hi-C.

MOTIVATION: High-resolution Hi-C data, capable of detecting chromatin features below the level of Topologically Associating Domains (TADs), significantly enhance our understanding of gene regulation. Micro-C, a variant of Hi-C incorporating a micrococcal nuclease (MNase) digestion step to examine interactions between nucleosome pairs, has been developed to overcome the resolution limitations of Hi-C. However, Micro-C experiments pose greater technical challenges compared to Hi-C, owing to the need for precise MNase digestion control and higher-resolution sequencing. Therefore, developing computational methods to derive Micro-C data from existing Hi-C datasets could lead to better usage of a large amount of existing Hi-C data in the scientific community and cost savings. RESULTS: We developed C2c ("high" or upper case C to "micro" or lower case c), a computational tool based on a residual neural network to learn the mapping between Hi-C and Micro-C contact matrices and then predict Micro-C contact matrices based on Hi-C contact matrices. Our evaluation results show that the predicted Micro-C contact matrices reveal more chromatin loops than the input Hi-C contact matrices, and more of the loops detected from predicted Micro-C match the promoter-enhancer interactions. Furthermore, we found that the mutual loops from real and predicted Micro-C better match the ChIA-PET data compared to Hi-C and real Micro-C loops, and the predicted Micro-C leads to more TAD-boundaries detected compared to the Hi-C data. The website URL of C2c can be found in the Data Availability Statement.

Three-dimensional chromatin mapping of sensory neurons reveals that cohesin-dependent genomic domains are required for axonal regeneration.

The in vivo three-dimensional genomic architecture of adult mature neurons at homeostasis and after medically relevant perturbations such as axonal injury remains elusive. Here we address this knowledge gap by mapping the three-dimensional chromatin architecture and gene expression programme at homeostasis and after sciatic nerve injury in wild-type and cohesin-deficient mouse sensory dorsal root ganglia neurons via combinatorial Hi-C and RNA-seq. We find that cohesin is required for the full induction of the regenerative transcriptional program, by organising 3D genomic domains required for the activation of regenerative genes. Importantly, loss of cohesin results in disruption of chromatin architecture at regenerative genes and severely impaired nerve regeneration. Together, these data provide an original three-dimensional chromatin map of adult sensory neurons in vivo and demonstrate a role for cohesin-dependent chromatin interactions in neuronal regeneration.

The Bioinformatic Applications of Hi-C and Linked Reads.

Long-range sequencing grants insight into additional genetic information beyond that which can be accessed by both short reads and modern long-read technology. Several new sequencing technologies are available for long-range datasets such as "Hi-C" and "Linked Reads" with high-throughput and high-resolution genome analysis, and are rapidly advancing the field of genome assembly, genome scaffolding, and more comprehensive variant identification. In this article, we focused on five major long-range sequencing technologies: high-throughput chromosome conformation capture (Hi-C), 10x Genomics Linked Reads, haplotagging, transposase enzyme linked long-read sequencing (TELL-seq), and single tube long fragment read (stLFR). We detailed the mechanisms and data products of the five platforms, introduced several of the most important applications, evaluated the quality of sequencing data from different platforms, and discussed the currently available bioinformatics tools. We hope this work will benefit the selection of appropriate long-range technology for specific biological studies.

HiCDiff: single-cell Hi-C data denoising with diffusion models.

The genome-wide single-cell chromosome conformation capture technique, i.e. single-cell Hi-C (ScHi-C), was recently developed to interrogate the conformation of the genome of individual cells. However, single-cell Hi-C data are much sparser than bulk Hi-C data of a population of cells, and noise in single-cell Hi-C makes it difficult to apply and analyze them in biological research. Here, we developed the first generative diffusion models (HiCDiff) to denoise single-cell Hi-C data in the form of chromosomal contact matrices. HiCDiff uses a deep residual network to remove the noise in the reverse process of diffusion and can be trained in both unsupervised and supervised learning modes. Benchmarked on several single-cell Hi-C test datasets, the diffusion models substantially remove the noise in single-cell Hi-C data. The unsupervised HiCDiff outperforms most supervised non-diffusion deep learning methods and achieves the performance comparable to the state-of-the-art supervised deep learning method in terms of multiple metrics, demonstrating that diffusion models are a useful approach to denoising single-cell Hi-C data. Moreover, its good performance holds on denoising bulk Hi-C data.

First Chromosome-Level Genome Assembly of a Ribbon Worm from the Hoplonemertea Clade, Emplectonema gracile, and Its Structural Annotation.

Genome-wide information has so far been unavailable for ribbon worms of the clade Hoplonemertea, the most species-rich class within the phylum Nemertea. While species within Pilidiophora, the sister clade of Hoplonemertea, possess a pilidium larval stage and lack stylets on their proboscis, Hoplonemertea species have a planuliform larva and are armed with stylets employed for the injection of toxins into their prey. To further compare these developmental, physiological, and behavioral differences from a genomic perspective, the availability of a reference genome for a Hoplonemertea species is crucial. Such data will be highly useful for future investigations toward a better understanding of molecular ecology, venom evolution, and regeneration not only in Nemertea but also in other marine invertebrate phyla. To this end, we herein present the annotated chromosome-level genome assembly for Emplectonema gracile (Nemertea; Hoplonemertea; Monostilifera; Emplectonematidae), an easily collected nemertean well suited for laboratory experimentation. The genome has an assembly size of 157.9 Mb. Hi-C scaffolding yielded chromosome-level scaffolds, with a scaffold N50 of 10.0 Mb and a score of 95.1% for complete BUSCO genes found as a single copy. Annotation predicted 20,684 protein-coding genes. The high-quality reference genome reaches an Earth BioGenome standard level of 7.C.Q50.

A haplotype-resolved reference genome of a long-distance migratory bat, Pipistrellus nathusii (Keyserling & Blasius, 1839).

We present a complete, chromosome-scale reference genome for the long-distance migratory bat Pipistrellus nathusii. The genome encompasses both haplotypic sets of autosomes and the separation of both sex chromosomes by utilizing highly accurate long-reads and preserving long-range phasing information through the use of three-dimensional chromatin conformation capture sequencing (Hi-C). This genome, accompanied by a comprehensive protein-coding sequence annotation, provides a valuable genomic resource for future investigations into the genomic bases of long-distance migratory flight in bats as well as uncovering the genetic architecture, population structure and evolutionary history of Pipistrellus nathusii. The reference-quality genome presented here gives a fundamental resource to further our understanding of bat genetics and evolution, adding to the growing number of high-quality genetic resources in this field. Here, we demonstrate its use in the phylogenetic reconstruction of the order Chiroptera, and in particular, we present the resources to allow detailed investigations into the genetic drivers and adaptations related to long-distance migration.

SMC-mediated dosage compensation in C. elegans evolved in the presence of an ancestral nematode mechanism.

Mechanisms of X chromosome dosage compensation have been studied extensively in a few model species representing clades of shared sex chromosome ancestry. However, the diversity within each clade as a function of sex chromosome evolution is largely unknown. Here, we anchor ourselves to the nematode Caenorhabditis elegans, for which a well-studied mechanism of dosage compensation occurs through a specialized structural maintenance of chromosomes (SMC) complex, and explore the diversity of dosage compensation in the surrounding phylogeny of nematodes. Through phylogenetic analysis of the C. elegans dosage compensation complex and a survey of its epigenetic signatures, including X-specific topologically associating domains (TADs) and X-enrichment of H4K20me1, we found that the condensin-mediated mechanism evolved recently in the lineage leading to Caenorhabditis through an SMC-4 duplication. Intriguingly, an independent duplication of SMC-4 and the presence of X-specific TADs in Pristionchus pacificus suggest that condensin-mediated dosage compensation arose more than once. mRNA-seq analyses of gene expression in several nematode species indicate that dosage compensation itself is ancestral, as expected from the ancient XO sex determination system. Indicative of the ancestral mechanism, H4K20me1 is enriched on the X chromosomes in Oscheius tipulae, which does not contain X-specific TADs or SMC-4 paralogs. Together, our results indicate that the dosage compensation system in C. elegans is surprisingly new, and condensin may have been co-opted repeatedly in nematodes, suggesting that the process of evolving a chromosome-wide gene regulatory mechanism for dosage compensation is constrained. Significance statement: X chromosome dosage compensation mechanisms evolved in response to Y chromosome degeneration during sex chromosome evolution. However, establishment of dosage compensation is not an endpoint. As sex chromosomes change, dosage compensation strategies may have also changed. In this study, we performed phylogenetic and epigenomic analyses surrounding Caenorhabditis elegans and found that the condensin-mediated dosage compensation mechanism in C. elegans is surprisingly new, and has evolved in the presence of an ancestral mechanism. Intriguingly, condensin-based dosage compensation may have evolved more than once in the nematode lineage, the other time in Pristionchus. Together, our work highlights a previously unappreciated diversity of dosage compensation mechanisms within a clade, and suggests constraints in evolving new mechanisms in the presence of an existing one.

Comparative 3D genome analysis between neural retina and retinal pigment epithelium reveals differential cis-regulatory interactions at retinal disease loci.

BACKGROUND: Vision depends on the interplay between photoreceptor cells of the neural retina and the underlying retinal pigment epithelium (RPE). Most genes involved in inherited retinal diseases display specific spatiotemporal expression within these interconnected retinal components through the local recruitment of cis-regulatory elements (CREs) in 3D nuclear space. RESULTS: To understand the role of differential chromatin architecture in establishing tissue-specific expression at inherited retinal disease loci, we mapped genome-wide chromatin interactions using in situ Hi-C and H3K4me3 HiChIP on neural retina and RPE/choroid from human adult donor eyes. We observed chromatin looping between active promoters and 32,425 and 8060 candidate CREs in the neural retina and RPE/choroid, respectively. A comparative 3D genome analysis between these two retinal tissues revealed that 56% of 290 known inherited retinal disease genes were marked by differential chromatin interactions. One of these was ABCA4, which is implicated in the most common autosomal recessive inherited retinal disease. We zoomed in on retina- and RPE-specific cis-regulatory interactions at the ABCA4 locus using high-resolution UMI-4C. Integration with bulk and single-cell epigenomic datasets and in vivo enhancer assays in zebrafish revealed tissue-specific CREs interacting with ABCA4. CONCLUSIONS: Through comparative 3D genome mapping, based on genome-wide, promoter-centric, and locus-specific assays of human neural retina and RPE, we have shown that gene regulation at key inherited retinal disease loci is likely mediated by tissue-specific chromatin interactions. These findings do not only provide insight into tissue-specific regulatory landscapes at retinal disease loci, but also delineate the search space for non-coding genomic variation underlying unsolved inherited retinal diseases.

From compartments to loops: understanding the unique chromatin organization in neuronal cells.

The three-dimensional organization of the genome plays a central role in the regulation of cellular functions, particularly in the human brain. This review explores the intricacies of chromatin organization, highlighting the distinct structural patterns observed between neuronal and non-neuronal brain cells. We integrate findings from recent studies to elucidate the characteristics of various levels of chromatin organization, from differential compartmentalization and topologically associating domains (TADs) to chromatin loop formation. By defining the unique chromatin landscapes of neuronal and non-neuronal brain cells, these distinct structures contribute to the regulation of gene expression specific to each cell type. In particular, we discuss potential functional implications of unique neuronal chromatin organization characteristics, such as weaker compartmentalization, neuron-specific TAD boundaries enriched with active histone marks, and an increased number of chromatin loops. Additionally, we explore the role of Polycomb group (PcG) proteins in shaping cell-type-specific chromatin patterns. This review further emphasizes the impact of variations in chromatin architecture between neuronal and non-neuronal cells on brain development and the onset of neurological disorders. It highlights the need for further research to elucidate the details of chromatin organization in the human brain in order to unravel the complexities of brain function and the genetic mechanisms underlying neurological disorders. This research will help bridge a significant gap in our comprehension of the interplay between chromatin structure and cell functions.

Removing unwanted variation between samples in Hi-C experiments.

Hi-C data are commonly normalized using single sample processing methods, with focus on comparisons between regions within a given contact map. Here, we aim to compare contact maps across different samples. We demonstrate that unwanted variation, of likely technical origin, is present in Hi-C data with replicates from different individuals, and that properties of this unwanted variation change across the contact map. We present band-wise normalization and batch correction, a method for normalization and batch correction of Hi-C data and show that it substantially improves comparisons across samples, including in a quantitative trait loci analysis as well as differential enrichment across cell types.