Lytic and Latent Genetic Diversity of the Epstein-Barr Virus Reveals Raji-Related Variants from Southeastern Brazil Associated with Recombination Markers.

Epstein-Barr virus (EBV) is a ubiquitous gammaherpesvirus etiologically associated with benign and malignant diseases. Since the pathogenic mechanisms of EBV are not fully understood, understanding EBV genetic diversity is an ongoing goal. Therefore, the present work describes the genetic diversity of the lytic gene BZLF1 in a sampling of 70 EBV-positive cases from southeastern Brazil. Additionally, together with the genetic regions previously characterized, the aim of the present study was to determine the impact of viral genetic factors that may influence EBV genetic diversity. Accordingly, the phylogenetic analysis of the BZLF1 indicated two main clades with high support, BZ-A and BZ-B (PP > 0.85). Thus, the BZ-A clade was the most diverse clade associated with the main polymorphisms investigated, including the haplotype Type 1 + V3 (p < 0.001). Furthermore, the multigene phylogenetic analysis (MLA) between BZLF1 and the oncogene LMP1 showed specific clusters, revealing haplotypic segregation that previous single-gene phylogenies from both genes failed to demonstrate. Surprisingly, the LMP1 Raji-related variant clusters were shown to be more diverse, associated with BZ-A/B and the Type 2/1 + V3 haplotypes. Finally, due to the high haplotypic diversity of the Raji-related variants, the number of DNA recombination-inducing motifs (DRIMs) was evaluated within the different clusters defined by the MLA. Similarly, the haplotype BZ-A + Raji was shown to harbor a greater number of DRIMs (p < 0.001). These results call attention to the high haplotype diversity of EBV in southeast Brazil and strengthen the hypothesis of the recombinant potential of South American Raji-related variants via the LMP1 oncogene.

Association of IL-18 gene polymorphisms with clinical aspects of hyperlipidemia in middle-aged and early people in the community.

Hyperlipidemia is notorious for causing coronary artery disease (CAD). IL-18 is a proinflammtory cytokine that contributes to the pathogenesis of CAD. Previous reports have revealed that genetic polymorphism of IL-18 is associated with its expression level as well as the susceptibility to CAD. In the present study, we aim to investigate the relationship between IL-18 single nucleotide polymorphisms (SNPs) and hyperlipidemia in the Han Chinese population in Taiwan. A total of 580 participants older than 30 were recruited from the community. We collected the demographics, self-reported disease histories, and lifestyles. We also assessed the levels of lipid profiles including total cholesterol (CHOL), triglyceride, low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol. Two SNPs, rs3882891C/A (intron 5) and rs1946518A/C (promoter -607) of IL-18 were elucidated by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods. Our results revealed that rs3882891 AA was associated with lower risk of hypercholesterolemia, higher CHOL and LDL-C in subjects (p=0.003, p=0.000 and p=0.005 separately), and rs1946518 CC was associated with hypercholesterolemia, higher CHOL and LDL-C as well (p=0.021, p=0.003 and p=0.001 separately) Furthermore, both SNPs were associated with IL-18 expression level, which was examined by Genotype-Tissue Expression (GTEx) Portal (p=0.042 and 0.016 separately). Finally, the haplotype of IL-18 was subsequently arranged in the order of rs3882891 and rs1946518. The result revealed that the AC haplotype of 2 IL-18 SNPs was also associated with lower risk of hypercholesterolemia, lower levels of CHOL and LDL-C (p=0.01, p=0.001 and 0.003). The current study is the first to report the association between IL-18 SNPs and hyperlipidemia in the Chinese Han population.

Association between Polymorphisms of Heat Shock Protein HSPA5 and Risk of Type 2 Diabetes Mellitus.

We studied the relationship between the HSPA5 gene polymorphisms and the risk of type 2 diabetes mellitus. Genotyping of three SNPs of the HSPA5 gene was performed in 1579 patients with type 2 diabetes mellitus and 1650 healthy individuals. It was found that the genotypes rs55736103-T/T, rs12009-G/G, and rs391957-T/C-T/T are associated with increased risk of type 2 diabetes in females. A rare haplotype, rs55736103C-rs12009A-rs391957T HSPA5, associated with a reduced risk of type 2 diabetes in females was found. Associations between polymorphisms of the HSPA5 gene encoding heat shock protein and the risk of type 2 diabetes mellitus were established for the first time.

Pangenome characterization and analysis of the NAC gene family reveals genes for Sclerotinia sclerotiorum resistance in sunflower (Helianthus annuus).

BACKGROUND: Sunflower (Helianthus annuus) is one of the most important economic crops in oilseed production worldwide. The different cultivars exhibit variability in their resistance genes. The NAC transcription factor (TF) family plays diverse roles in plant development and stress responses. With the completion of the H. annuus genome sequence, the entire complement of genes coding for NACs has been identified. However, the reference genome of a single individual cannot cover all the genetic information of the species. RESULTS: Considering only a single reference genome to study gene families will miss many meaningful genes. A pangenome-wide survey and characterization of the NAC genes in sunflower species were conducted. In total, 139 HaNAC genes are identified, of which 114 are core and 25 are variable. Phylogenetic analysis of sunflower NAC proteins categorizes these proteins into 16 subgroups. 138 HaNACs are randomly distributed on 17 chromosomes. SNP-based haplotype analysis shows haplotype diversity of the HaNAC genes in wild accessions is richer than in landraces and modern cultivars. Ten HaNAC genes in the basal stalk rot (BSR) resistance quantitative trait loci (QTL) are found. A total of 26 HaNAC genes are differentially expressed in response to Sclerotinia head rot (SHR). A total of 137 HaNAC genes are annotated in Gene Ontology (GO) and are classified into 24 functional groups. GO functional enrichment analysis reveals that HaNAC genes are involved in various functions of the biological process. CONCLUSIONS: We identified NAC genes in H. annuus (HaNAC) on a pangenome-wide scale and analyzed S. sclerotiorum resistance-related NACs. This study provided a theoretical basis for further genomic improvement targeting resistance-related NAC genes in sunflowers.

Ancestry of the major long-range regulatory site of the α-globin genes in the Portuguese population with the common 3.7 kb α-thalassemia deletion.

BACKGROUND: The α-Major Regulatory Element (α-MRE), also known as HS-40, is located upstream of the α-globin gene cluster and has a crucial role in the long-range regulation of the α-globin gene expression. This enhancer is polymorphic and several haplotypes were identified in different populations, with haplotype D almost exclusively found in African populations. The purpose of this research was to identify the HS-40 haplotype associated with the 3.7 kb α-thalassemia deletion (-α3.7del) in the Portuguese population, and determine its ancestry and influence on patients' hematological phenotype. METHODS AND RESULTS: We selected 111 Portuguese individuals previously analyzed by Gap-PCR to detect the presence of the -α3.7del: 50 without the -α3.7del, 34 heterozygous and 27 homozygous for the -α3.7del. The HS-40 region was amplified by PCR followed by Sanger sequencing. Four HS-40 haplotypes were found (A to D). The distribution of HS-40 haplotypes and genotypes are significantly different between individuals with and without the -α3.7del, being haplotype D and genotype AD the most prevalent in patients with this deletion in homozygosity. Furthermore, multiple correspondence analysis revealed that individuals without the -α3.7del are grouped with other European populations, while samples with the -α3.7del are separated from these and found more closely related to the African population. CONCLUSION: This study revealed for the first time an association of the HS-40 haplotype D with the -α3.7del in the Portuguese population, and its likely African ancestry. These results may have clinical importance as in vitro analysis of haplotype D showed a decrease in its enhancer activity on α-globin gene.

Chromosome-Y haplogroups in Asturias (Northern Spain) and their association with severe COVID-19.

The main objective of this study was to determine whether the common Y-haplogroups were be associated with the risk of developing severe COVID-19 in Spanish male. We studied 479 patients who required hospitalization due to COVID-19 and 285 population controls from the region of Asturias (northern Spain), They were genotyped for several polymorphisms that define the common European Y-haplogroups. We compared the frequencies between patients and controls aged ≤ 65 and >65 years. There were no different haplogroup frequencies between the two age groups of controls. Haplogroup R1b was less common in patients aged ≤65 years. Haplogroup I was more common in the two patient´s groups compared to controls (p = 0.02). Haplogroup R1b was significantly more frequent among hypertensive patients, without difference between the hypertensive and normotensive controls. This suggested that R1b could increase the risk for severe COVID-19 among male with pre-existing hypertension. In conclusion, we described the Y-haplogroup structure among Asturians. We found an increased risk of severe COVID-19 among haplogroup I carriers, and a significantly higher frequency of R1b among hypertensive patients. These results indicate that Y-chromosome variants could serve as markers to define the risk of developing a severe form of COVID-19.

Comprehensive genetic analysis of facioscapulohumeral muscular dystrophy by Nanopore long-read whole-genome sequencing.

BACKGROUND: Facioscapulohumeral muscular dystrophy (FSHD) is a high-prevalence autosomal dominant neuromuscular disease characterized by significant clinical and genetic heterogeneity. Genetic diagnosis of FSHD remains a challenge because it cannot be detected by standard sequencing methods and requires a complex diagnosis workflow. METHODS: We developed a comprehensive genetic FSHD detection method based on Oxford Nanopore Technologies (ONT) whole-genome sequencing. Using a case-control design, we applied this procedure to 29 samples and compared the results with those from optical genome mapping (OGM), bisulfite sequencing (BSS), and whole-exome sequencing (WES). RESULTS: Using our ONT-based method, we identified 59 haplotypes (35 4qA and 24 4qB) among the 29 samples (including a mosaic sample), as well as the number of D4Z4 repeat units (RUs). The pathogenetic D4Z4 RU contraction identified by our ONT-based method showed 100% concordance with OGM results. The methylation levels of the most distal D4Z4 RU and the double homeobox 4 gene (DUX4) detected by ONT sequencing are highly consistent with the BSS results and showed excellent diagnostic efficiency. Additionally, our ONT-based method provided an independent methylation profile analysis of two permissive 4qA alleles, reflecting a more accurate scenario than traditional BSS. The ONT-based method detected 17 variations in three FSHD2-related genes from nine samples, showing 100% concordance with WES. CONCLUSIONS: Our ONT-based FSHD detection method is a comprehensive method for identifying pathogenetic D4Z4 RU contractions, methylation level alterations, allele-specific methylation of two 4qA haplotypes, and variations in FSHD2-related genes, which will all greatly improve genetic testing for FSHD.

KSNP: a fast de Bruijn graph-based haplotyping tool approaching data-in time cost.

Long reads that cover more variants per read raise opportunities for accurate haplotype construction, whereas the genotype errors of single nucleotide polymorphisms pose great computational challenges for haplotyping tools. Here we introduce KSNP, an efficient haplotype construction tool based on the de Bruijn graph (DBG). KSNP leverages the ability of DBG in handling high-throughput erroneous reads to tackle the challenges. Compared to other notable tools in this field, KSNP achieves at least 5-fold speedup while producing comparable haplotype results. The time required for assembling human haplotypes is reduced to nearly the data-in time.

Environmental DNA unveils deep phylogeographic structure of a freshwater fish.

Phylogeography bears an important part in ecology and evolution. However, current phylogeographic studies are largely constrained by limited numbers of individual samples. Using an environmental DNA (eDNA) assay for phylogeographic analyses, this study provides detailed information regarding the history of Siberian stone loach Barbatula toni, a primary freshwater fish across the whole range of Hokkaido, Japan. Based on an eDNA metabarcoding on 293 river water samples, we detected eDNA from B. toni in 189 rivers. A total of 51 samples, representing the entire island, were then selected from the B. toni eDNA-positive sample set for the subsequent analyses. To elucidate the phylogeographic structure of B. toni, newly developed eDNA metabarcoding primers (Barba-cytb-F/R) were applied to these samples, specifically targeting their haplotypic variation in cytochrome b. After a bioinformatic processing to mitigate haplotypic false positives, a total of 50 eDNA haplotypes were identified. Two regionally restricted, genetically distinct lineages of the species were revealed as a result of phylogeographic analyses on the haplotypes and tissue-derived DNA from B. toni. According to a molecular clock analysis, they have been genetically isolated for at least 1.5 million years, suggesting their ancient origin and colonisation of Hokkaido, presumably in the glacial periods. These results demonstrate how freshwater fishes can alter their distributions over evolutionary timescales and how eDNA assay can deepen our understanding of phylogeography.

Haplotype-resolved assembly of diploid and polyploid genomes using quantum computing.

Precision medicine's emphasis on individual genetic variants highlights the importance of haplotype-resolved assembly, a computational challenge in bioinformatics given its combinatorial nature. While classical algorithms have made strides in addressing this issue, the potential of quantum computing remains largely untapped. Here, we present the vehicle routing problem (VRP) assembler: an approach that transforms this task into a vehicle routing problem, an optimization formulation solvable on a quantum computer. We demonstrate its potential and feasibility through a proof of concept on short synthetic diploid and triploid genomes using a D-Wave quantum annealer. To tackle larger-scale assembly problems, we integrate the VRP assembler with Google's OR-Tools, achieving a haplotype-resolved local assembly across the human major histocompatibility complex (MHC) region. Our results show encouraging performance compared to Hifiasm with phasing accuracy approaching the theoretical limit, underscoring the promising future of quantum computing in bioinformatics.

Association between CBL gene polymorphism and susceptibility of microscopic polyangiitis in a Chinese population: A case-control analysis.

OBJECTIVE: To assess whether Casitas B-lineage lymphoma (CBL) gene polymorphism influences the risk of microscopic polyangiitis (MPA) in Chinese populations. METHODS: In total, 266 MPA patients and 297 healthy controls were recruited for a case-control study. Five CBL SNPs were genotyped using multiplex polymerase chain reaction and high-throughput sequencing. The relationship between SNPs and the risk of MPA under different genetic models was evaluated by SNPstats. SNP-SNP interaction was analyzed by generalized multifactor dimensionality reduction (GMDR). Finally, the association between CBL SNPs and treatment effects were assessed. RESULTS: The results showed that CBL rs2276083 was associated with decreasing MPA risk under dominant (OR: 0.53; p = 0.014) and recessive models (OR: 0.52; p = 0.0034). Stratification analysis indicated that rs2276083 and rs2509671 in age < 60 years, rs2276083 in female or in Han population were protective factors for MPA. The CBL haplotype (A-A-G-C-T) was associated with an increased risk of MPA. GMDR suggested that CBL rs2276083, phosphatidylinositol-4, 5-bisphosphate 3-kinase catalytic subunit alpha (PI3KCA) rs1607237, and autophagy-related gene 7 (ATG7) rs7549008 might interact with each other in MPA development (p = 0.0107). CBL rs1047417 with AG genotype and rs11217234 with AG genotype had better clinical treatment effects than other two genotypes (p = 0.048 and p = 0.025, respectively). CONCLUSION: The genetic polymorphism of CBL had a potential association with the risk of MPA and clinical treatment effects in Guangxi population in China.

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.

The 5' untranslated region variant rs3811050 C/T of the interleukin-38 encoding gene is associated with susceptibility to rheumatoid arthritis in Iraqi women.

BACKGROUND: Interleukin (IL)-38, the latest member of the IL-1 cytokine family, is proposed to have a pathogenic role in rheumatoid arthritis (RA). It is encoded by the IL1F10 gene, which harbors single nucleotide polymorphisms (SNPs) that may predict the risk of autoimmune diseases. Among them are 5' untranslated region (UTR) SNPs, which play a key role in post-transcriptional control, but have not been studied in Iraqi RA patients. METHODS: Two novel IL1F10 5'UTR SNPs (rs3811050 C/T and rs3811051 T/G) were explored in RA and control women (n = 120 and 110, respectively). SNPs were genotyped using TaqMan assay. An ELISA kit was used to measure serum IL-38 concentrations. RESULTS: A reduced risk of RA was associated with rs3811050 T allele and CT genotype (corrected probability [pc] = 0.01 and < 0.001, respectively), while there was no significant association with rs3811051. Haplotype analysis demonstrated that C-T haplotype was associated with a 1.65-fold greater risk of RA, whereas a reduced risk was linked to T-G haplotype. IL-38 concentrations were higher in patients than in controls (p < 0.001). In addition, IL-38 showed acceptable performance in distinguishing between RA and control women (p < 0.001). When IL-38 concentrations were stratified according to SNP genotypes, no significant differences were found. CONCLUSIONS: The rs3811050 variant was more likely to affect RA susceptibility in Iraqi women, and the T allele may play a role in reducing disease risk. IL-38 concentrations were elevated in RA patients, but were not affected by the rs3811050 and rs3811051 genotypes.

Evolutionary characteristics of the mitochondrial NADH dehydrogenase subunit 6 gene in some populations of four sympatric Mustela species (Mustelidae, Mammalia) from central Europe.

BACKGROUND: Selection on or reticulate evolution of mtDNA is documented in various mammalian taxa and could lead to misleading phylogenetic conclusions if not recognized. We sequenced the MT-ND6 gene of four sympatric Mustelid species of the genus Mustela from some central European populations. We hypothesised positive selection on MT-ND6, given its functional importance and the different body sizes and life histories of the species, even though climatic differences may be unimportant for adaptation in sympatry. METHODS AND RESULTS: MT-ND6 genes were sequenced in 187 sympatric specimens of weasels, Mustela nivalis, stoats, M. erminea, polecats, M. putorius, and steppe polecats, M. eversmannii, from eastern Austria and of fourteen allopatric polecats from eastern-central Germany. Median joining networks, neighbour joining and maximum likelihood analyses as well as Bayesian inference grouped all species according to earlier published phylogenetic models. However, polecats and steppe polecats, two very closely related species, shared the same two haplotypes. We found only negative selection within the Mustela sequences, including 131 downloaded ones covering thirteen species. Positive selection was observed on three MT-ND6 codons of other mustelid genera retrieved from GenBank. CONCLUSIONS: Negative selection for MT-ND6 within the genus Mustela suggests absence of both environmental and species-specific effects of cellular energy metabolism despite large species-specific differences in body size. The presently found shared polymorphism in European polecats and steppe polecats may result from ancestral polymorphism before speciation and historical or recent introgressive hybridization; it may indicate mtDNA capture of steppe polecats by M. putorius in Europe.

Unraveling the Pathogenetic Mechanisms Underlying the Association between Specific Mitochondrial DNA Haplogroups and Parkinson's Disease.

Variants of mitochondrial DNA (mtDNA) have been identified as risk factors for the development of Parkinson's disease (PD). However, the underlying pathogenetic mechanisms remain unclear. Cybrid models carrying various genotypes of mtDNA variants were tested for resistance to PD-simulating MPP+ treatment. The most resistant line was selected for transcriptome profiling, revealing specific genes potentially influencing the resistant characteristic. We then conducted protein validation and molecular biological studies to validate the related pathways as the influential factor. Cybrids carrying the W3 mtDNA haplogroup demonstrated the most resistance to the MPP+ treatment. In the transcriptome study, PPP1R15A was identified, while further study noted elevated expressions of the coding protein GADD34 across all cybrids. In the study of GADD34-related mitochondrial unfolding protein response (mtUPR), we found that canonical mtUPR, launched by the phosphate eIF2a, is involved in the resistant characteristic of specific mtDNA to MPP+ treatment. Our study suggests that a lower expression of GADD34 in the late phase of mtUPR may prolong the mtUPR process, thereby benefitting protein homeostasis and facilitating cellular resistance to PD development. We herein demonstrate that GADD34 plays an important role in PD development and should be further investigated as a target for the development of therapies for PD.

Delimiting Species with Single-Locus DNA Sequences.

DNA sequences are increasingly used for large-scale biodiversity inventories. Because these genetic data avoid the time-consuming initial sorting of specimens based on their phenotypic attributes, they have been recently incorporated into taxonomic workflows for overlooked and diverse taxa. Major statistical developments have accompanied this new practice, and several models have been proposed to delimit species with single-locus DNA sequences. However, proposed approaches to date make different assumptions regarding taxon lineage history, leading to strong discordance whenever comparisons are made among methods. Distance-based methods, such as Automatic Barcode Gap Discovery (ABGD) and Assemble Species by Automatic Partitioning (ASAP), rely on the detection of a barcode gap (i.e., the lack of overlap in the distributions of intraspecific and interspecific genetic distances) and the associated threshold in genetic distances. Network-based methods, as exemplified by the REfined Single Linkage (RESL) algorithm for the generation of Barcode Index Numbers (BINs), use connectivity statistics to hierarchically cluster-related haplotypes into molecular operational taxonomic units (MOTUs) which serve as species proxies. Tree-based methods, including Poisson Tree Processes (PTP) and the General Mixed Yule Coalescent (GMYC), fit statistical models to phylogenetic trees by maximum likelihood or Bayesian frameworks.Multiple webservers and stand-alone versions of these methods are now available, complicating decision-making regarding the most appropriate approach to use for a given taxon of interest. For instance, tree-based methods require an initial phylogenetic reconstruction, and multiple options are now available for this purpose such as RAxML and BEAST. Across all examined species delimitation methods, judicious parameter setting is paramount, as different model parameterizations can lead to differing conclusions. The objective of this chapter is to guide users step-by-step through all the procedures involved for each of these methods, while aggregating all necessary information required to conduct these analyses. The "Materials" section details how to prepare and format input files, including options to align sequences and conduct tree reconstruction with Maximum Likelihood and Bayesian inference. The Methods section presents the procedure and options available to conduct species delimitation analyses, including distance-, network-, and tree-based models. Finally, limits and future developments are discussed in the Notes section. Most importantly, species delimitation methods discussed herein are categorized based on five indicators: reliability, availability, scalability, understandability, and usability, all of which are fundamental properties needed for any approach to gain unanimous adoption within the DNA barcoding community moving forward.

The effects of ANRIL polymorphisms on colorectal cancer, tumor stage, and tumor grade among Iranian population.

BACKGROUND: Colorectal cancer (CRC) is a type of neoplasm, developing in the colon or rectum. The exact etiology of CRC is not well known, but the role of genetic, epigenetic, and environmental factors are established in its pathogenesis. Therefore, the aim of this research was to explore the effects of ANRIL polymorphisms on the CRC and its clinical findings. METHODS AND RESULTS: The peripheral blood specimens were collected from 142 CRC patients and 225 controls referred to Milad Hospital, Tehran, Iran. PCR- RFLP method was used to analyze ANRIL rs1333040, rs10757274 rs4977574, and rs1333048 polymorphisms. The ANRIL rs1333040 polymorphism was related to a higher risk of CRC in the co-dominant, dominant, and log-additive models. ANRIL rs10757274, rs4977574, and rs1333048 polymorphisms showed no effect on CRC susceptibility. The CGAA and TGGA haplotypes of ANRIL rs1333040/ rs10757274/ rs4977574/rs1333048 polymorphisms were associated with the higher and the lower risk of CRC respectively. The rs1333040 polymorphism was associated with higher TNM stages (III and IV). The frequency of ANRIL rs10757274 polymorphism was lower in CRC patients over 50 years of age only in the dominant model. In addition, the rs10757274 was associated with well differentiation in CRC patients. CONCLUSION: The ANRIL rs1333040 polymorphism was associated with a higher risk of CRC and higher TNM stages. ANRIL rs10757274 polymorphism was associated with the well-differentiated tumor in CRC.

Haplotype-resolved assembly of auto-polyploid genomes via combining Hi-C and gametic data.

Haplotype-resolved genome assembly plays a crucial role in understanding allele-specific functions. However, obtaining haplotype-resolved assembly for auto-polyploid genomes remains challenging. Existing methods can be classified into reference-based phasing, assembly-based phasing, and gamete binning. Nevertheless, there is a lack of cost-effective and efficient methods for haplotyping auto-polyploid genomes. In this study, we propose a novel phasing algorithm called PolyGH, which combines Hi-C and gametic data. We conducted experiments on tetraploid potato cultivars and divided the method into three steps. Firstly, gametic data was utilized to bin non-collapsed contigs, followed by merging adjacent fragments of the same type within the same contig. Secondly, accurate Hi-C signals related to differential genomic regions were acquired using unique k-mers. Finally, collapsed fragments were assigned to haplotigs based on combined Hi-C and gametic signals. Comparing PolyGH with Hi-C-based and gametic data-based methods, we found that PolyGH exhibited superior performance in haplotyping auto-polyploid genomes when integrating both data types. This approach has the potential to enhance haplotype-resolved assembly for auto-polyploid genomes.

HaplotagLR: An efficient and configurable utility for haplotagging long reads.

Understanding the functional effects of sequence variation is crucial in genomics. Individual human genomes contain millions of variants that contribute to phenotypic variability and disease risks at the population level. Because variants rarely act in isolation, we must consider potential interactions of neighboring variants to accurately predict functional effects. We can accomplish this using haplotagging, which matches sequencing reads to their parental haplotypes using alleles observed at known heterozygous variants. However, few published tools for haplotagging exist and these share several technical and usability-related shortcomings that limit applicability, in particular a lack of insight or control over error rates, and lack of key metrics on the underlying sources of haplotagging error. Here we present HaplotagLR: a user-friendly tool that haplotags long sequencing reads based on a multinomial model and existing phased variant lists. HaplotagLR is user-configurable and includes a basic error model to control the empirical FDR in its output. We show that HaplotagLR outperforms the leading haplotagging method in simulated datasets, especially at high levels of specificity, and displays 7% greater sensitivity in haplotagging real data. HaplotagLR advances both the immediate utility of haplotagging and paves the way for further improvements to this important method.

Nanopore sequencing with T2T-CHM13 for accurate detection and preventing the transmission of structural rearrangements in highly repetitive heterochromatin regions in human embryos.

BACKGROUND: Structural rearrangements in highly repetitive heterochromatin regions can result in miscarriage or foetal malformations; however, detecting and preventing the transmission of these rearrangements has been challenging. Recently, the completion of sequencing of the complete human genome (T2T-CHM13) has made it possible to accurately characterise structural rearrangements in these regions. We developed a method based on T2T-CHM13 and nanopore sequencing to detect and block structural rearrangements in highly repetitive heterochromatin sequences. METHODS: T2T-CHM13-based "Mapping Allele with Resolved Carrier Status" was performed for couples who carry structural rearrangements in heterochromatin regions. Using nanopore sequencing and the T2T-CHM13 reference genome, the precise breakpoints of inversions and translocations close to the centromere were detected and haplotypes were constructed using flanking single-nucleotide polymorphisms (SNPs). Haplotype linkage analysis was then performed by comparing consistent parental SNPs with embryonic SNPs to determine whether the embryos carried hereditary inversions or balanced translocations. Based on copy number variation and haplotype linkage analysis, we transplanted normal embryos, which were further verified by an amniotic fluid test. RESULTS: To validate this approach, we used nanopore sequencing of families with inversions and reciprocal translocations close to the centromere. Using the T2T-CHM13 reference genome, we accurately detected inversions and translocations in centromeres, constructed haplotypes and prevented the transmission of structural rearrangements in the offspring. CONCLUSIONS: This study represents the first successful application of T2T-CHM13 in human reproduction and provides a feasible protocol for detecting and preventing the transmission of structural rearrangements of heterochromatin in embryos.