Linkage disequilibrium and heritability of copy-number polymorphisms within duplicated regions of the human genome

Studies of copy-number variation and linkage disequilibrium (LD) have typically excluded complex regions of the genome that are rich in duplications and prone to rearrangement. In an attempt to assess the heritability and LD of copy-number polymorphisms (CNPs) in duplication-rich regions of the genome, we profiled copy-number variation in 130 putative "rearrangement hotspot regions" among 269 individuals of European, Yoruba, Chinese, and Japanese ancestry analyzed by the International HapMap Consortium. Eighty-four hotspot regions, corresponding to 257 bacterial artificial chromosome (BAC) probes, showed evidence of copy-number differences. Despite a predisposing genetic architecture, no polymorphism was ever observed in the remaining 46 "rearrangement hotspots," and we suggest these represent excellent candidate sites for pathogenic rearrangements. We used a combination of BAC-based and high-density customized oligonucleotide arrays to resolve the molecular basis of structural rearrangements. For common variants (frequency >10%), we observed a distinct bias against copy-number losses, suggesting that deletions are subject to purifying selection. Heritability estimates did not differ significantly from 1.0 among the majority (30 of 34) of loci analyzed, consistent with normal Mendelian inheritance. Some of the CNPs in duplication-rich regions showed strong LD with nearby single-nucleotide polymorphisms (SNPs) and were observed to segregate on ancestral SNP haplotypes. However, LD with the best available SNP markers was weaker than has been reported for deletion polymorphisms in less complex regions of the genome. These observations may be accounted for by a low density of SNP data in duplicated regions, challenges in mapping and typing the CNPs, and the possibility that CNPs in these regions have rearranged on multiple haplotype backgrounds. Our results underscore the need for complete maps of genetic variation in duplication-rich regions of the genome.     

A Bayesian approach to copy-number-polymorphism analysis in nuclear pedigrees

Segmental copy-number polymorphisms (CNPs) represent a significant component of human genetic variation and are likely to contribute to disease susceptibility. These potentially multiallelic and highly polymorphic systems present new challenges to family-based genetic-analysis tools that commonly assume codominant markers and allow for no genotyping error. The copy-number quantitation (CNP phenotype) represents the total number of segmental copies present in an individual and provides a means to infer, rather than to observe, the underlying allele segregation. We present an integrated approach to meet these challenges, in the form of a graphical model in which we infer the underlying CNP phenotype from the (single or replicate) quantitative measure within the analysis while assuming an allele-based system segregating through the pedigree. This approach can be readily applied to the study of any form of genetic measure, and the construction permits extension to a wide variety of hypothesis tests. We have implemented the basic model for use with nuclear families, and we illustrate its application through an analysis of the CNP located in gene CCL3L1 in 201 families with asthma.     

Molecular methods for genotyping complex copy number polymorphisms

Genome structural variation shows remarkable complexity with respect to copy number, sequence content and distribution. While the discovery of copy number polymorphisms (CNP) has increased exponentially in recent years, the transition from discovery to genotyping has proved challenging, particularly for CNPs embedded in complex regions of the genome. CNPs that are collectively common in the population and possess a dynamic range of copy numbers have proved the most difficult to genotype in association studies. This is in some part due to technical limitations of genotyping assays and the sequence properties of the genomic region being analyzed. Here we describe in detail the basis of a number of molecular techniques used to genotype complex CNPs, compare and contrast these approaches for determination of multi-allelic copy number, and discuss the potential application of these techniques in genetic studies.     

CNstream: A method for the identification and genotyping of copy number polymorphisms using Illumina microarrays

Background  

Understanding the genetic basis of disease risk in depth requires an exhaustive knowledge of the types of genetic variation. Very recently, Copy Number Variants (CNVs) have received much attention because of their potential implication in common disease susceptibility. Copy Number Polymorphisms (CNPs) are of interest as they segregate at an appreciable frequency in the general population (i.e. > 1%) and are potentially implicated in the genetic basis of common diseases.     

Genome-Wide Association Study Identified CNP12587 Region Underlying Height Variation in Chinese Females

Introduction

Human height is a highly heritable trait considered as an important factor for health. There has been limited success in identifying the genetic factors underlying height variation. We aim to identify sequence variants associated with adult height by a genome-wide association study of copy number variants (CNVs) in Chinese.

Methods

Genome-wide CNV association analyses were conducted in 1,625 unrelated Chinese adults and sex specific subgroup for height variation, respectively. Height was measured with a stadiometer. Affymetrix SNP6.0 genotyping platform was used to identify copy number polymorphisms (CNPs). We constructed a genomic map containing 1,009 CNPs in Chinese individuals and performed a genome-wide association study of CNPs with height.

Results

We detected 10 significant association signals for height (p<0.05) in the whole population, 9 and 11 association signals for Chinese female and male population, respectively. A copy number polymorphism ({"type":"entrez-protein","attrs":{"text":"CNP12587","term_id":"893597589","term_text":"CNP12587"}}CNP12587, chr18:54081842-54086942, p = 2.41×10⁻⁴) was found to be significantly associated with height variation in Chinese females even after strict Bonferroni correction (p = 0.048). Confirmatory real time PCR experiments lent further support for CNV validation. Compared to female subjects with two copies of the CNP, carriers of three copies had an average of 8.1% decrease in height. An important candidate gene, ubiquitin-protein ligase NEDD4-like (NEDD4L), was detected at this region, which plays important roles in bone metabolism by binding to bone formation regulators.

Conclusions

Our findings suggest the important genetic variants underlying height variation in Chinese.     

Patient-specific induced pluripotent stem cells as“disease-in-adish”models for inherited cardiomyopathies and channelopathies–15 years of research

Among inherited cardiac conditions,a special place is kept by cardiomyopathies(CMPs)and channelopathies(CNPs),which pose a substantial healthcare burden due to the complexity of the therapeutic management and cause early mortality.Like other inherited cardiac conditions,genetic CMPs and CNPs exhibit incomplete penetrance and variable expressivity even within carriers of the same pathogenic deoxyribonucleic acid variant,challenging our understanding of the underlying pathogenic mechanisms.Until recently,the lack of accurate physiological preclinical models hindered the investigation of fundamental cellular and molecular mechanisms.The advent of induced pluripotent stem cell(iPSC)technology,along with advances in gene editing,offered unprecedented opportunities to explore hereditary CMPs and CNPs.Hallmark features of iPSCs include the ability to differentiate into unlimited numbers of cells from any of the three germ layers,genetic identity with the subject from whom they were derived,and ease of gene editing,all of which were used to generate“disease-in-a-dish”models of monogenic cardiac conditions.Functionally,iPSC-derived cardiomyocytes that faithfully recapitulate the patient-specific phenotype,allowed the study of disease mechanisms in an individual-/allele-specific manner,as well as the customization of therapeutic regimen.This review provides a synopsis of the most important iPSC-based models of CMPs and CNPs and the potential use for modeling disease mechanisms,personalized therapy and deoxyribonucleic acid variant functional annotation.     

Effect of different sampling strategies for a single geographic region in Yemen on standard genetic analyses of mitochondrial DNA sequence data

Collection of biological samples is the foundation of genetic studies ranging from estimation of genetic diversity to reconstruction of population history. Sample collections are intended to accurately represent the genetic, biological, ecological, cultural, geographic, and/or linguistic diversity of a particular region or population by providing a small, but representative, set of samples. In this study, we analyze human mitochondrial DNA variation in samples collected using four different sampling strategies to represent the same geographic region. Specifically, samples were collected from a village, a rural area, a regional clinic, and a national university in the governorate of Dhamar in Yemen. All samples were assayed for mitochondrial hypervariable region I DNA sequence variation and data were subjected to standard molecular genetic analyses. Our results suggest that analyses in which individual DNA sequences are explicitly compared or evaluated, e.g. phylogenetic and network analyses, may be more sensitive to sample collection design than analyses in which data are averaged across individuals or are analyzed more indirectly, e.g. summary statistics.     

Environmental Heterogeneity Explains the Genetic Structure of Continental and Mediterranean Populations of Fraxinus angustifolia Vahl

Tree species with wide distributions often exhibit different levels of genetic structuring correlated to their environment. However, understanding how environmental heterogeneity influences genetic variation is difficult because the effects of gene flow, drift and selection are confounded. We investigated the genetic variation and its ecological correlates in a wind-pollinated Mediterranean tree species, Fraxinus angustifolia Vahl, within a recognised glacial refugium in Croatia. We sampled 11 populations from environmentally divergent habitats within the Continental and Mediterranean biogeographical regions. We combined genetic data analyses based on nuclear microsatellite loci, multivariate statistics on environmental data and ecological niche modelling (ENM). We identified a geographic structure with a high genetic diversity and low differentiation in the Continental region, which contrasted with the significantly lower genetic diversity and higher population divergence in the Mediterranean region. The positive and significant correlation between environmental and genetic distances after controlling for geographic distance suggests an important influence of ecological divergence of the sites in shaping genetic variation. The ENM provided support for niche differentiation between the populations from the Continental and Mediterranean regions, suggesting that contemporary populations may represent two divergent ecotypes. Ecotype differentiation was also supported by multivariate environmental and genetic distance analyses. Our results suggest that despite extensive gene flow in continental areas, long-term stability of heterogeneous environments have likely promoted genetic divergence of ashes in this region and can explain the present-day genetic variation patterns of these ancient populations.     

Number of individuals and molecular markers to use in genetic differentiation studies

Molecular markers are frequently used to study genetic variation among individuals within or between populations. Differences in marker banding patterns can be used to verify if individuals do, or do not, represent distinct groups or populations. Only in 2005, more than 500 studies used molecular markers to group individuals in clusters. Such studies make use of an arbitrary number of molecular markers from each of an arbitrary number of individuals presumed to represent distinct genotypes. However, the greater the genetic variation, the more likely a larger number of individuals and markers will be needed to capture a population's genetic signature. The numbers of both, markers and individuals included thus affect the way in which individuals are organized through cluster analyses, thereby affecting the conclusions drawn. Here we present a method that provides statistical criteria to verify that individual and marker sample sizes are sufficient to accurately depict genetic differentiation among different populations. Our method uses a resampling technique to assess the reproducibility of obtaining a particular grouping pattern for specific data sets. It thus, allows to estimate the robustness of the results obtained without including additional individuals, or markers.     

Determining the source of individuals: multilocus genotyping in nonequilibrium population genetics

Recently founded populations represent an enormous challenge for genetic analysis: new populations are often genetically impoverished, making it hard to find sufficiently variable markers, and what little variation is present tends to be ancestral, rendering phylogenetic methods inappropriate. Recently, novel genetic markers and new statistical analyses have made multilocus genotyping an invaluable tool in the fledgling field of nonequilibrium population genetics. Such advances are not of mere academic interest but address questions of great economic, medical and conservation significance.     

Morphometrics Parallel Genetics in a Newly Discovered and Endangered Taxon of Galápagos Tortoise

Galápagos tortoises represent the only surviving lineage of giant tortoises that exhibit two different types of shell morphology. The taxonomy of Galápagos tortoises was initially based mainly on diagnostic morphological characters of the shell, but has been clarified by molecular studies indicating that most islands harbor monophyletic lineages, with the exception of Isabela and Santa Cruz. On Santa Cruz there is strong genetic differentiation between the two tortoise populations (Cerro Fatal and La Reserva) exhibiting domed shell morphology. Here we integrate nuclear microsatellite and mitochondrial data with statistical analyses of shell shape morphology to evaluate whether the genetic distinction and variability of the two domed tortoise populations is paralleled by differences in shell shape. Based on our results, morphometric analyses support the genetic distinction of the two populations and also reveal that the level of genetic variation is associated with morphological shell shape variation in both populations. The Cerro Fatal population possesses lower levels of morphological and genetic variation compared to the La Reserva population. Because the turtle shell is a complex heritable trait, our results suggest that, for the Cerro Fatal population, non-neutral loci have probably experienced a parallel decrease in variability as that observed for the genetic data.     

Differential gene flow of mitochondrial and nuclear DNA markers among chromosomal races of Australian morabine grasshoppers (Vandiemenella, viatica species group)

Recent theoretical developments have led to a renewed interest in the potential role of chromosomal rearrangements in speciation. Australian morabine grasshoppers (genus Vandiemenella, viatica species group) provide an excellent study system to test this potential role of chromosomal rearrangements because they show extensive chromosomal variation and formed the basis of a classic chromosomal speciation model. There are three chromosomal races, viatica19, viatica17, and P24(XY), on Kangaroo Island, South Australia, forming five parapatric populations with four putative contact zones among them. We investigate the extent to which chromosomal variation among these populations may be associated with barriers to gene flow. Population genetic and phylogeographical analyses using 15 variable allozyme loci and the elongation factor-1alpha (EF-1alpha) gene indicate that the three races represent genetically distinct taxa. In contrast, analyses of the mitochondrial cytochrome c oxidase subunit I (COI) gene show the presence of three distinctive and geographically localized groups that do not correspond with the distribution of the chromosomal races. These discordant population genetic patterns are likely to result from introgressive hybridization between the chromosomal races and range expansions/contractions. Overall, these results suggest that reduction of nuclear gene flow may be associated with chromosomal variation, or underlying genetic variation linked with chromosomal variation, whereas mitochondrial gene flow appears to be independent of this variation in these morabine grasshoppers. The identification of an intact contact zone between P24(XY) and viatica17 offers considerable potential for further investigation of molecular mechanisms that maintain distinct nuclear genomes among the chromosomal races.     

PEGylated ceria nanoparticles used for radioprotection on human liver cells under γ-ray irradiation

Ceria nanoparticles (CNPs) have recently been shown to protect cells and animals from radiation-induced damage. However, most of the CNPs used in previous studies were either naked or weakly protected by surfactants, which inevitably encounter many obstacles in biological applications. Here, alendronate was used as an ideal anchor to graft polyethylene glycol (PEG) onto CNPs, leading to enhanced stability, reduced cytotoxicity, and improved biological properties. Further investigation assessed the protective ability of the nanoparticles against radiation-induced effects for human normal liver cells (L-02), indicating that the PEGylated CNPs (CNPs–AL–PEG) were more efficient than naked CNPs. We determined that enhanced Ce³⁺/Ce⁴⁺ ratios improved intracellular dispersion and that the ameliorated intracellular distribution of CNPs–AL–PEG contributes to the elevated expression of SOD2, which leads to increased protection of normal cells against ROS and reduces the oxidatively generated DNA damage. These studies hold tremendous promise for radioprotection and biological applications.     

Global population genetic structure and male-mediated gene flow in the green sea turtle (Chelonia mydas): analysis of microsatellite loci

We assessed the degree of population subdivision among global populations of green sea turtles, Chelonia mydas, using four microsatellite loci. Previously, a single-copy nuclear DNA study indicated significant male-mediated gene flow among populations alternately fixed for different mitochondrial DNA haplotypes and that genetic divergence between populations in the Atlantic and Pacific Oceans was more common than subdivisions among populations within ocean basins. Even so, overall levels of variation at single-copy loci were low and inferences were limited. Here, the markedly more variable microsatellite loci confirm the presence of male-mediated gene flow among populations within ocean basins. This analysis generally confirms the genetic divergence between the Atlantic and Pacific. As with the previous study, phylogenetic analyses of genetic distances based on the microsatellite loci indicate a close genetic relationship among eastern Atlantic and Indian Ocean populations. Unlike the single-copy study, however, the results here cannot be attributed to an artifact of general low variability and likely represent recent or ongoing migration between ocean basins. Sequence analyses of regions flanking the microsatellite repeat reveal considerable amounts of cryptic variation and homoplasy and significantly aid in our understanding of population connectivity. Assessment of the allele frequency distributions indicates that at least some of the loci may not be evolving by the stepwise mutation model.     

The perils of single gene trees — mitochondrial versus single-copy nuclear DNA variation in white-eyes (Aves: Zosteropidae)

Phylogenetic relationships among animal populations and species commonly have been inferred from patterns of variation observed within a single gene system, most often the mitochondrial genome. Analysis of restriction site variation in the mitochondrial DNA of two species of white-eye ( Zosterops lateralis and Z. lutea ) in Australia produced a single gene tree that does not accurately represent the organismal tree. In contrast, patterns of variation at two anonymous, single-copy nuclear DNA loci revealed a phylogeography consistent with traditional classification of the species. Discordance between mitochondrial DNA and single-copy nuclear DNA variation is probably the result of past hybridization between Z. lateralis and Z. lutea , evidence of which has been lost from the nuclear genome by recombination. This study provides a clear empirical demonstration that single gene genealogies cannot be assumed to accurately represent the true phylogenies, and emphasizes the need for composite genetic analyses.     

Population-genetic properties of differentiated human copy-number polymorphisms

Copy-number variants (CNVs) can reach appreciable frequencies in the human population, and recent discoveries have shown that several of these copy-number polymorphisms (CNPs) are associated with human diseases, including lupus, psoriasis, Crohn disease, and obesity. Despite new advances, significant biases remain in terms of CNP discovery and genotyping. We developed a method based on single-channel intensity data and benchmarked against copy numbers determined from sequencing read depth to successfully obtain CNP genotypes for 1495 CNPs from 487 human DNA samples of diverse ethnic backgrounds. This microarray contained CNPs in segmental duplication-rich regions and insertions of sequences not represented in the reference genome assembly or on standard SNP microarray platforms. We observe that CNPs in segmental duplications are more likely to be population differentiated than CNPs in unique regions (p = 0.015) and that biallelic CNPs show greater stratification when compared to frequency-matched SNPs (p = 0.0026). Although biallelic CNPs show a strong correlation of copy number with flanking SNP genotypes, the majority of multicopy CNPs do not (40% with r > 0.8). We selected a subset of CNPs for further characterization in 1876 additional samples from 62 populations; this revealed striking population-differentiated structural variants in genes of clinical significance such as OCLN, a tight junction protein involved in hepatitis C viral entry. Our microarray design allows these variants to be rapidly tested for disease association and our results suggest that CNPs (especially those that cannot be imputed from SNP genotypes) might have contributed disproportionately to human diversity and selection.     

Partitioning of copy-number genotypes in pedigrees

Background  

Copy number variations (CNVs) and polymorphisms (CNPs) have only recently gained the genetic community's attention. Conservative estimates have shown that CNVs and CNPs might affect more than 10% of the genome and that they may be at least as important as single nucleotide polymorphisms in assessing human variability. Widely used tools for CNP analysis have been implemented in Birdsuite and PLINK for the purpose of conducting genetic association studies based on the unpartitioned total number of CNP copies provided by the intensities from Affymetrix's Genome-Wide Human SNP Array. Here, we are interested in partitioning copy number variations and polymorphisms in extended pedigrees for the purpose of linkage analysis on familial data.     

Various physicochemical and surface properties controlling the bioactivity of cerium oxide nanoparticles

Amidst numerous emerging nanoparticles, cerium oxide nanoparticles (CNPs) possess fascinating pharmacological potential as they can be used as a therapeutic for various oxidative stress-associated chronic diseases such as cancer, inflammation and neurodegeneration due to unique redox cycling between Ce³⁺ and Ce⁴⁺ oxidation states on their surface. Lattice defects generated by the formation of Ce³⁺ ions and compensation by oxygen vacancies on CNPs surface has led to switching between CeO₂ and CeO_2–x during redox reactions making CNPs a lucrative catalytic nanoparticle capable of mimicking key natural antioxidant enzymes such as superoxide dismutase and catalase. Eventually, most of the reactive oxygen species and nitrogen species in biological system are scavenged by CNPs via an auto-regenerative mechanism in which a minimum dose can exhibit catalytic activity for a longer duration. Due to the controversial outcomes on CNPs toxicity, considerable attention has recently been drawn towards establishing relationships between the physicochemical properties of CNPs obtained by different synthesis methods and biological effects ranging from toxicity to therapeutics. Unlike non-redox active nanoparticles, variations in physicochemical properties and the surface properties of CNPs obtained from different synthesis methods can significantly affect their biological activity (inactive, antioxidant, or pro-oxidant). Moreover, these properties can influence the biological identity, cellular interactions, cellular uptake, biodistribution, and therapeutic efficiency. This review aims to highlight the critical role of various physicochemical and the surface properties of CNPs controlling their biological activity based on 165 cited references.     

Do seed transfer zones for ecological restoration reflect the spatial genetic variation of the common grassland species Lathyrus pratensis?

A common ecological restoration approach is the reestablishment of vegetation using seed mixtures. To preserve the natural genetic pattern of plant species local seed material should be used. Consequently, seed transfer zones (seed production areas and seed provenance regions) have been delineated for ecological restoration in Germany. Although it is assumed that these transfer zones represent genetic variation, there remains a lack of empirical data. In this study, we analyzed whether seed transfer zones reflect the genetic variation of the common grassland species Lathyrus pratensis. We sampled 706 individuals from 37 populations in Bavaria, Germany and analyzed genetic variation using amplified fragment length polymorphisms. In our study, we observed higher levels of genetic variation and fragment rarity in the southern Bavarian populations compared to northern populations. Our analyses revealed a strong genetic differentiation between southern and northern Bavarian populations delineated along the Danube River. However, seed production areas and seed provenance regions reflected genetic variation of L. pratensis only to a limited degree. Our study illustrates that the level of genetic variation within populations strongly depends on population history. Furthermore, the geomorphological and climatic attributes, which have been used to delineate seed provenance regions, do not reduce gene flow among populations. Seed collections for gene banks and seed production should comprise seeds from populations in southern and northern Bavaria representing the strong genetic variation between both regions, but prioritize southern populations due to higher levels of variation.     

Fluorescent carbon nanoparticles obtained from charcoal via green methods and their application for sensing Fe3+ in an aqueous medium

Two green methods (microwave and hydrothermal) were employed for the preparation of water dispersible fluorescent carbon nanoparticles (CNPs) from activated charcoal. Microwave and hydrothermally synthesized carbon nanoparticles, (MW‐CNPs) and (HT‐CNPs), respectively were characterized by microscopic and spectroscopic techniques. A detailed study of their fluorescence characteristics was made. MW‐CNPs and HT‐CNPs were tested for metal ion selectivity in aqueous medium. MW‐CNPs showed selectivity for Fe³⁺ among the tested metal ions and important studies such as for interference, linear range and limit of detection were carried out. The application of MW‐CNPs for detection of Fe³⁺ in water was demonstrated.