Biology subjects

Zhang, M.

Publications and source records attributed to Zhang, M..

At least 397 records · Page 22Linked to original sources

bioRxiv2023

High-throughput single-cell sorting by stimulated Raman-activated cell ejection

Single-cell sorting is essential to explore cellular heterogeneity in biology and medicine. Recently developed Raman-activated cell sorting (RACS) circumvents the limitations of fluorescence-activated cell sorting, such as the cytotoxicity of labels. However, the sorting throughputs of all forms of RACS are limited by the intrinsically small cross-section of spontaneous Raman scattering. Here, we report a stimulated Raman-activated cell ejection (S-RACE) platform that enables high-throughput single-cell sorting based on high-resolution multi-channel stimulated Raman chemical imaging, in situ image decomposition, and laser-induced cell ejection. The performance of this platform was illustrated by sorting a mixture of 1 m polymer beads, where 95% yield, 98% purity, and 14 events per second throughput were achieved. Notably, our platform allows live cell ejection, allowing for the growth of single colonies of bacteria and fungi after sorting. To further illustrate the chemical selectivity, lipid-rich Rhodotorula glutinis cells were successfully sorted from a mixture with Saccharomyces cerevisiae, confirmed by downstream quantitative PCR. Furthermore, by integrating a closed-loop feedback control circuit into the system, we realized real-time single-cell imaging and sorting, and applied this method to precisely eject regions of interest from a rat brain tissue section. The reported S-RACE platform opens exciting opportunities for a wide range of single-cell applications in biology and medicine. Significance statementImage-guided single-cell sorting is a potent tool in diverse biological applications. Current microfluidic cell sorting methods encounter challenges in handling smaller cells and are not applicable to tissue sections. To address these challenges, we have developed a stimulated Raman-activated cell ejection (S-RACE) platform, which is the first demonstration of single-cell ejection coupled with coherent Raman scattering. S-RACE allows label-free chemical imaging guided cell sorting through multispectral stimulated Raman scattering (SRS) imaging, on-the-fly image analysis, and laser-induced cell ejection. Versatile applications of S-RACE to a wide range of samples, such as polymer particles, single-live bacteria, single-live fungus, and tissue sections, are demonstrated.

bioengineering↗

bioRxiv2023

Distinctive and complementary roles of default mode network subsystems in semantic cognition

The default mode network (DMN) typically deactivates to external tasks, yet supports semantic cognition. It comprises medial temporal (MT), core, and fronto-temporal (FT) subsystems, but its functional organisation is unclear: the requirement for perceptual coupling versus decoupling, input modality (visual/verbal), type of information (social/spatial) and control demands all potentially affect its recruitment. We examined the effect of these factors on activation and deactivation of DMN subsystems during semantic cognition, across four task-based functional magnetic resonance imaging (fMRI) datasets, and localised these responses in whole-brain state space defined by gradients of intrinsic connectivity. FT showed activation consistent with a central role across domains, tasks and modalities, although it was most responsive to abstract, verbal tasks; this subsystem uniquely showed more tuned states characterised by increases in both activation and deactivation when semantic retrieval demands were higher. MT also activated to both perceptually-coupled (scenes) and decoupled (autobiographical memory) tasks, and showed stronger responses to picture associations, consistent with a role in scene construction. Core DMN consistently showed deactivation, especially to externally-oriented tasks. These diverse contributions of DMN subsystems to semantic cognition were related to their location on intrinsic connectivity gradients: activation was closer to sensory-motor cortex than deactivation, particularly for FT and MT, while activation for core DMN was distant from both visual cortex and cognitive control. These results reveal distinctive yet complementary DMN responses: MT and FT support different memory-based representations that are accessed externally and internally, while deactivation in core DMN is associated with demanding, external semantic tasks. Significance StatementWe delineate the functional organisation of DMN in semantic cognition, examining effects of perceptual coupling versus decoupling, input modality (visual/verbal), domain (social/spatial) and control demands across DMN subsystems in four fMRI datasets. These subsystems played complementary roles in semantic cognition related to their locations on gradients of intrinsic connectivity. Medial temporal and frontotemporal subsystems supported visuospatial and abstract conceptual information respectively, across both internally and externally-focussed tasks, while deactivation in core DMN was associated with focussed and externally-oriented semantic states. We conclude that both content and process are relevant to the functional architecture of DMN in semantic cognition.

neuroscience↗

bioRxiv2023

Default mode network shows distinct emotional and contextual responses yet common effects of retrieval demands across tasks

The default mode network (DMN) lies towards the heteromodal end of the principal gradient of intrinsic connectivity, maximally separated from sensory-motor cortex. It supports memory-based cognition, including the capacity to retrieve conceptual and evaluative information from sensory inputs, and to generate meaningful states internally; however, the functional organisation of DMN that can support these distinct modes of retrieval remains unclear. We used fMRI to examine whether activation within subsystems of DMN differed as a function of retrieval demands, or the type of information to be retrieved, or both. In a picture association task, participants retrieved two types of semantic features about contexts and emotions: in the generate phase, these associations were retrieved from a novel picture, while in a switch phase, participants retrieved a new association for the same image. Semantic context and emotion trials were associated with dissociable DMN subnetworks, indicating that a key dimension of DMN organisation relates to the type of information being accessed. The fronto-temporal and medial temporal DMN showed a preference for emotional and contextual associations, respectively. Relative to the generate phase, the switch phase recruited clusters closer to the heteromodal apex of the principal gradient - a cortical hierarchy separating unimodal and heteromodal regions. There were no differences in this effect between association types. Instead, memory switching was associated with a distinct subnetwork associated with controlled internal cognition. These findings delineate distinct patterns of DMN recruitment for different kinds of associations yet common responses across tasks that reflect retrieval demands. Key pointsO_LIRetrieval of contextual and emotional features relies on distinct default mode subnetworks. C_LIO_LINovelty of visual input has an equivalent effect across default mode subnetworks. C_LIO_LISecondary associations retrieved from re-presented pictures activate heteromodal end of the principal gradient. C_LI

neuroscience↗

bioRxiv2023

Mechanism of ZNF143 in chromatin looping and 3D genome organization

The transcription factor ZNF143 contains a central domain of seven zinc fingers in a tandem array and is involved in 3D genome construction; however, the mechanism by which ZNF143 functions in chromatin looping remains unclear. Here, we show that ZNF143 directionally recognizes a diverse range of genomic sites within enhancers and promoters and is required for chromatin looping between these sites. In addition, ZNF143 is located between CTCF and cohesin at numerous CTCF sites, and ZNF143 removal narrows the space between CTCF and cohesin. Moreover, genetic deletion of ZNF143, in conjunction with acute CTCF depletion, reveals that ZNF143 and CTCF collaborate to regulate higher-order chromatin organization. Thus, ZNF143 is recruited by CTCF to the CTCF sites to regulate TAD formation, whereas directional recognition of genomic DNA motifs directly by ZNF143 itself regulates promoter activity via chromatin looping.

genetics↗

bioRxiv2023

Adaptive Functions of Structural Variants in Human Brain Development

Quantifying the structural variants (SVs) in nonhuman primates could provide a niche to clarify the genetic backgrounds underlying human-specific traits, but such resource is largely lacking. Here, we report an accurate SV atlas in a population of 562 rhesus macaques, verified by two public SV benchmarks, an inhouse benchmark of eight macaque genomes with long-read sequencing and another inhouse benchmark of one macaque genome with whole-genome assembly. This accurate, quantitative SV map indicates stronger purifying selection on inversions, one type of poorly-clarified SVs to date, especially for those located on regulatory regions, suggesting a strategy for prioritizing inversions with the most important functions. Based on the distribution and the evolutionary features of these inversions in macaque population, we then identified 75 human-specific inversions, clarified their functional effects and prioritized them. Notably, the top-ranked inversions have substantially shaped the human transcriptome, through their dual-effects of reconfiguring the ancestral genomic architecture and introducing regional mutation hotspots at the inverted regions. As a proof-of-concept, we linked APCDD1, located on one of these inversions with the highest rank score and downregulated in human brains, to neuronal maturation. The accumulation of human-specific mutations on its promoter region, accelerated by the formation of the inversion, contributed to the decreased expression in humans. Notably, the overexpression of APCDD1 could accelerate the neuronal maturation, while its depletion in mice delays the neuronal maturation. This study thus highlights the contribution of SVs, especially the inversions, to the distinct features in human brain development.

evolutionary biology↗

bioRxiv2023

Perisaccadic and Attentional Remapping of Receptive Fields in Lateral Intraparietal Area and Frontal Eye Fields

The nature and function of perisaccadic receptive-field (RF) remapping have been controversial. We used a delayed saccade task to reduce previous confounds and examined the remapping time course in areas LIP and FEF. In the delay period, the RF shift direction turned from the initial fixation to the saccade target. In the perisaccadic period, RFs first shifted toward the target (convergent remapping) but around the time of saccade onset/offset, the shifts became predominantly toward the post-saccadic RF locations (forward remapping). Thus, unlike forward remapping that depends on the corollary discharge (CD) of the saccade command, convergent remapping appeared to follow attention from the initial fixation to the target. We modelled the data with attention-modulated and CD-gated connections, and showed that both sets of connections emerged automatically in neural networks trained to update stimulus retinal locations across saccades. Our work thus unifies previous findings into a mechanism for transsaccadic visual stability.

neuroscience↗

bioRxiv2023

Reconciling ASPP-p53 Binding Mode Discrepancies through an Ensemble Binding Framework that Bridges Crystallography and NMR Data

ASPP2 and iASPP bind to p53 through their conserved ANK-SH3 domains to respectively promote and inhibit p53-dependent cell apoptosis. While crystallography has indicated that these two proteins employ distinct surfaces of their ANK-SH3 domains to bind to p53, solution NMR data has suggested similar surfaces. In this study, we employed multi-scale molecular dynamics (MD) combined with free energy calculations to reconcile the discrepancy in the binding modes. We demonstrated that the binding mode based solely on a single crystal structure does not enable iASPPs RT loop to engage with p53s C-terminal linker--a verified interaction. Instead, an ensemble of simulated iASPP-p53 complexes facilitates this interaction. We showed that the ensemble-average inter-protein contacting residues and NMR-detected interfacial residues align well with ASPP proteins, and the ensemble-average binding free energies better match experimental Kd values compared to single crystallgarphy-determined binding mode. For iASPP, the sampled ensemble complexes can be grouped into two classes, resembling the binding modes determined by crystallography and solution NMR. We thus propose that crystal packing shifts the equilibrium of binding modes towards the crystallographydetermined one. Lastly, we show that the ensemble binding complexes are sensitive to p53s intrinsically disordered regions (IDRs), attesting to experimental observations that these IDRs contribute to biological functions. Our results provide a dynamic and ensemble perspective for scrutinizing these important cancer-related protein-protein interactions (PPIs).

biophysics↗

bioRxiv2023

A novel MARV glycoprotein-specific antibody with potentials of broad-spectrum neutralization to filovirus

Marburg virus (MARV) is one of the filovirus species that cause deadly hemorrhagic fever in humans, with mortality rates up to 90%. Neutralizing antibodies represent ideal candidates to prevent or treat virus disease. However, no antibody has been approved for MARV treatment to date. In this study, we identified a novel human antibody named AF-03 that targeted MARV glycoprotein (GP). AF-03 possessed a high binding affinity to MARV GP and showed neutralizing and protective activities against the pseudotyped MARV in vitro and in vivo. Epitope identification, including molecular docking and experiment-based analysis of mutated species, revealed that AF-03 recognized the Niemann-Pick C1 (NPC1) binding domain within GP1. Interestingly, we found the neutralizing activity of AF-03 to pseudotyped Ebola viruses (EBOV, SUDV, and BDBV) harboring cleaved GP instead of full-length GP. Furthermore, NPC2-fused AF-03 exhibited neutralizing activity to several filovirus species and EBOV mutants via binding to CI-MPR. In conclusion, this work demonstrates that AF-03 represents a promising therapeutic cargo for filovirus-caused disease.

immunology↗

bioRxiv2023

Sde Proteins Coordinate Ubiquitin Utilization and Phosphoribosylation to Promote Establishment and Maintenance of the Legionella Replication Vacuole

The Legionella pneumophila Sde family of translocated proteins promotes host tubular endoplasmic reticulum (ER) rearrangements that are tightly linked to phosphoribosyl-ubiquitin (pR-Ub) modification of Reticulon 4 (Rtn4). Sde proteins have two additional activities of unclear relevance to the infection process: K63 linkage-specific deubiquitination and phosphoribosyl modification of polyubiquitin (pR-Ub). We show here that the deubiquitination activity (DUB) stimulates ER rearrangements while pR-Ub protects the replication vacuole from cytosolic surveillance by autophagy. Loss of DUB activity is tightly linked to lowered pR-Ub modification of Rtn4, consistent with the DUB activity fueling the production of pR-Ub-Rtn4. In parallel, phosphoribosyl modification of polyUb, in a region of the protein known as the isoleucine patch, prevents binding by the autophagy adapter p62. An inability of Sde mutants to modify polyUb results in immediate p62 association, a critical precursor to autophagic attack. The ability of Sde WT to block p62 association decays quickly after bacterial infection, as predicted by the presence of previously characterized L. pneumophila effectors that inactivate Sde and remove polyUb. In sum, these results show that the accessory Sde activities act to stimulate ER rearrangements and protect from host innate immune sensing in a temporal fashion.

microbiology↗

bioRxiv2023

Genome report: Whole genome sequence and annotation of Penstemon davidsonii

Penstemon is the most speciose flowering plant genus endemic to North America. Penstemon species diverse morphology and adaptation to various environments have made them a valuable model system for studying evolution, but the absence of publicly available reference genomes limits possible research directions. Here we report the first reference genome assembly and annotation for Penstemon davidsonii. Using PacBio long-read sequencing and Hi-C scaffolding technology, we constructed a de novo reference genome of 437,568,744 bases, with a contig N50 of 40 Mb and L50 of 5. The annotation includes 18,199 gene models, and both the genome and transcriptome assembly contain over 95% complete eudicot BUSCOs. This genome assembly will serve as a valuable reference for studying the evolutionary history and genetic diversity of the Penstemon genus.

genomics↗

bioRxiv2023

CaMKII autophosphorylation but not downstream kinase activity is required for synaptic memory

CaMKII plays a critical role in long-term potentiation (LTP), a well-established model for learning and memory through the enhancement of synaptic transmission. Biochemical studies indicate that CaMKII catalyzes a phosphotransferase (kinase) reaction of both itself (autophosphorylation) and of multiple downstream target proteins. However, whether either type of phosphorylation plays any role in the synaptic enhancing action of CaMKII remains hotly contested. We have designed a series of experiments to define the minimal requirements for the synaptic enhancement by CaMKII. We find that autophosphorylation of T286 and further binding of CaMKII to the GluN2B subunit are required both for initiating LTP and for its maintenance (synaptic memory). Once bound to the NMDA receptor, the synaptic action of CaMKII occurs in the absence of kinase activity. Thus, autophosphorylation, together with binding to the GluN2B subunit, are the only two requirements for CaMKII in synaptic memory.

neuroscience↗

bioRxiv2023

The role of neutrophil typing in homeostatic resilience protective model of inflammatory preconditioning

Sepsis is a systemic inflammatory response syndrome (SIRS) characterized by a dysregulated host response to infection that results in organ dysfunction. The response of neutrophils to early inflammation is critical, interferon regulatory factors 4 (IRF4) and interferon regulatory factors 5 (IRF5) are expressed in most cell types of the immune system, but the relationship between the role of IRF4 and IRF5 in preconditioning-protected septic peritonitis mouse model and neutrophils response remains unknown. In this study, we used an E.coli-induced septic peritonitis mouse model and zebrafish model to explore the relationship between neutrophil inflammation and IRF subtype changes in the process of septic peritonitis. Mechanistically, we found that the protein complex formed by polymorphonuclear neutrophils (PMN) and IRF5/MyD88 can secrete pro-inflammatory factors and directly kill invading bacteria. Author SummaryNumerous animal and clinical sepsis studies have shown that markedly impaired recruitment of neutrophils to sites of infection and failure to clear bacteria contribute to excessive inflammation and increased mortality in sepsis. In our previous work, we established an Escherichia coli (E.coli) lethal septic peritonitis model and a preconditioning-protected septic peritonitis mouse model. In the early stage of septic peritonitis, a large number of PMN infiltrate the peritoneal cavity, which produces an inflammatory response to invasive bacteria. In this study, the switching process between pro-inflammatory and anti-inflammatory attracted our attention. Our research shows that IRF5-IRF4 regulatory axis leads to a PMN phenotype switch in sepsis and the expression of IRF5 and IRF4 was related to PMN pro-inflammatory (N1 type) and anti-inflammatory (N2 type) phenotypes, respectively.

immunology↗

bioRxiv2023

Prokaryotic microbial diversity analysis and metabolic function prediction of salt lakes on the Qinghai-Tibet Plateau

The Dong Taijinar Salt Lake (DT) and Xi Taijinar Salt Lake (XT) have been widely studied as mineral-rich areas. However, little is known about the composition and distribution of the microbial communities in these two lakes. In this study, metagenomics sequencing was used to analyze the diversity and potential functions of the microbial communities in DT and XT. According to our report, the salinity of DT (332.18-358.30 g/L) was 10 times higher than that of XT (20.09-36.83 g/L). Interestingly, the dominant domain in DT was Archaea (96.16%), while that in XT was Bacteria (93.09%). The distribution of Bacteria in the DT revealed 33 phyla and 1717 genera. The dominant genus in DT was Marinobacillus, which was positively correlated with total phosphorus content. There were four main phyla and 153 genera identified in the Archaea of DT. The most abundant Archaea genera in DT were Natronomonas (24.61%) and Halorubrum (23.69%), which were mainly positively correlated with the Na+, Ca2+, and Cl- contents. Similarly, there were 33 phyla and 1906 genera of Bacteria in XT, and Loktanella was the dominant genus. The archaeal taxonomy in XT mainly included four phyla and 149 genera. Proteobacteria and Euryarchaeota were the most abundant bacterial and archaeal phyla in the two salt lakes. Analysis of the halophilic mechanisms of the microorganisms identified in these two salt lakes revealed that the Bacteria in XT preferred to synthesize compatible solutes, whereas the Archaea in DT preferred a "salt-in" adaptation strategy in salt-stressed environments. IMPORTANCEThe Qinghai-Tibet Plateau is the origin of many lakes and mountains in China. Among them, the Dong Taijinar and Xi Taijinar salt lakes are important biological resources with unknown microbial community compositions and functional potentials. The results of this study revealed significant differences in the distribution of Bacteria and Archaea between the two salt lakes. Salinity mainly drives lower biodiversity and restricted bacterial growth and metabolism in the high-salinity and near-saturated Dong Taijinar Salt Lake. This study not only identifies the key microorganisms in two penetrating salt lakes, but also provides insights into the mechanisms of salinity tolerance and the unknown ecological functions of microorganisms in extreme environments.

microbiology↗

bioRxiv2023

Single-cell analysis characterizes PLK1 as a promoter of immunosuppressive tumor microenvironment in LUAD

PLK1 (Polo-like kinase 1) plays a critical role in the progression of lung adenocarcinoma (LUAD). Recent studies have unveiled that targeting PLK1 improves the efficacy of immunotherapy, highlighting its important role in the regulation of tumor immunity. Nevertheless, our understanding of the intricate interplay between PLK1 and the tumor microenvironment (TME) remains incomplete. Here, using genetically engineered mouse model and single-cell RNA-seq analysis, we report that PLK1 promotes an immunosuppressive TME in LUAD, characterized with enhanced M2 polarization of tumor associated macrophages (TAM) and dampened antigen presentation process. Mechanistically, elevated PLK1 coincides with increased secretion of CXCL2 cytokine, which promotes M2 polarization of TAM and diminishes expression of class II major histocompatibility complex (MHC-II) in professional antigen-presenting cells. Furthermore, PLK1 negatively regulates MHC-II expression in cancer cells, which has been shown to be associated with compromised tumor immunity and unfavorable patient outcomes. Taken together, our results reveal PLK1 as a novel modulator of TME in LUAD and provide possible therapeutic interventions.

cancer biology↗

bioRxiv2023

Preserved stimulus representations in frontal cortex as a potential neural mechanism for flexible control in working memory

When holding visual information temporarily in working memory (WM), the neural representation of the memorandum is distributed across various cortical regions, including visual and frontal cortices. However, the role of stimulus representation in visual and frontal cortices during WM has been controversial. Here we tested the hypothesis that stimulus representation persists in the frontal cortex to facilitate flexible control demands in WM. During functional MRI, participants flexibly switched between simple WM maintenance of visual stimulus or more complex rule-based categorization of maintained stimulus on a trial-by-trial basis. Our results demonstrated enhanced stimulus representation in the frontal cortex that tracked demands for active WM control and enhanced stimulus representation in the visual cortex that tracked demands for precise WM maintenance. This differential frontal stimulus representation traded off with the newly-generated category representation with varying control demands. Simulation using multi-module recurrent neural networks replicated human neural patterns when stimulus information was preserved for network readout. Altogether, these findings help reconcile the long-standing debate in WM research, and provide empirical and computational evidence that flexible stimulus representation in the frontal cortex during WM serves as a potential neural coding scheme to accommodate the ever-changing environment.

neuroscience↗

bioRxiv2023

USP20 deubiquitinates and stabilizes the ER-phagy receptor FAM134B to drive ER-phagy

The endoplasmic reticulum (ER) serves as a hub for various essential cellular processes, and maintaining ER homeostasis is essential for cell function. ER-phagy is a selective process that removes impaired ER subdomains through autophagosomes and lysosomal degradation. While the involvement of ubiquitination in autophagy regulation is well-established, its role in ER-phagy remains unclear. In our study, we screened deubiquitinating enzymes involved in ER-phagy and identified USP20 (ubiquitin specific peptidase 20) as a key regulator of ER-phagy under stress conditions. USP20 specifically cleaves K63- and K48-linked ubiquitin chains on the ER-phagy receptor FAM134B/RETREG1 (reticulophagy regulator 1), thereby stabilizing the substrate and promoting ER-phagy. Remarkably, despite lacking a transmembrane domain, USP20 is recruited to the ER through its interaction with VAPs (vesicle-associated membrane proteins). VAPs facilitate the recruitment of early autophagy proteins, including WIPI2, to specific ER subdomains, where USP20 and FAM134B are enriched. This recruitment of WIPI2 and other proteins plays a crucial role in facilitating FAM134B-mediated ER-phagy in response to cellular stress. Our findings highlight the critical role of USP20 in maintaining ER homeostasis by deubiquitinating and stabilizing FAM134B at distinct ER subdomains, where USP20 further recruits VAPs and promotes efficient ER-phagy.

molecular biology↗

bioRxiv2023

Comprehensive catalogs for microbial genes and metagenome-assembled genomes of the swine lower respiratory tract microbiome identify the relationship of microbial species with lung lesions

Understanding the community structure and functional capacity of the lower respiratory tract microbiome is crucial for elucidating its roles in respiratory tract diseases. However, there are few studies about it owing to the difficulty in obtaining microbial samples from the lower respiratory tract. Here we collected 745 microbial samples from the porcine lower respiratory tract by harvesting 675 pigs, and constructed a gene catalog containing 10,337,194 nonredundant genes, of which only 30% could be annotated taxonomically. We obtained 397 metagenome-assembled genomes (MAGs) including 111 MAGs with high-quality. These 397 MAGs were further clustered into 292 species-level genome bins (SGBs), among which 56% SGBs are unknown with current databases. Combining with the lung lesion phenotype, we found that Mycoplasma hyopneumoniae strains and the adhesion-related virulence factors harboring in their genomes were significantly associated with porcine lung lesions, implying the role of adhesion and overgrowth of pathogenic M. hyopneumoniae in host lung diseases. This study provided important resources for the study of porcine lower respiratory tract microbiome and lung health.

microbiology↗

bioRxiv2023

T2T-YAO: a Telomere-to-telomere Assembled Diploid Reference Genome for Han Chinese

Since its initial release in 2001, the human reference genome has been continuously improved in both continuity and accuracy, and the recently-released telomere-to-telomere version--T2T-CHM13--reaches its top quality after 20 years of effort. However, T2T-CHM13 does not represent an authentic diploid human genome, but rather one derived from a simplified, nearly homozygous genome of a hydatidiform mole cell line. To address this limitation and provide an alternative pertinent to the Chinese population, the largest ethnic group in the world, we have assembled a complete diploid human genome of a male Han Chinese, T2T-YAO, which includes telomere-to-telomere assemblies for all the 22+X+M and 22+Y chromosomes in his two haploids inherited separately from his parents. Both haplotypes contain no artificial sequences or model nucleotides and possess a high quality comparable to CHM13, with fewer than one error per [~]14 Mb. Derived from the individual who lives in the aboriginal region of Han Chinese, T2T-YAO shows clear ancestry and potential genetic continuity from the ancient ancestors of the Han population. Each haplotype of T2T-YAO possesses [~]340 Mb exclusive sequences and [~]3100 unique genes as compared to CHM13, and their genome sequences show greater genetic distance to CHM13 than to each other in terms of nucleotide polymorphism and structural variations. The construction of T2T-YAO would serve as a high-quality diploid reference that enables precise delineation of genomic variations in a haplotype-sensitive manner, which could advance our understandings in human evolution, hereditability of diseases and phenotypes, especially within the context of the unique variations of the Chinese population.

genetics↗