Search Results - EGA European Genome-Phenome Archive

Whole Genome Sequencing of clonal expansions of single healthy somatic cells (human, female)

HSC73_clone: Bone marrow mononuclear cells from the healthy 73 years old female were thawed and labeled with Alexa-Fluor 488-conjugated anti-CD34 (581, Biolegend), Alexa-Fluor 700-conjugated anti-CD38 (HIT2, eBioscience), a cocktail of APC-conjugated lineage antibodies consisting of anti-CD4 (RPA-T4), anti-CD8 (RPA-T8), anti-CD11b (ICRF44), anti-CD20 (2H7), anti-CD56 (B159, all BD Biosciences), anti-CD14 (61D3), anti-CD19 (HIB19) and anti-CD235a (HIR2, all eBiocience) and 1 micro-gram/ml propidium iodide (Sigma). Using a BD FACSAria cell sorter, single Lin-CD34+CD38-PI- cells were individually sorted into low-adhesion 96-well tissue culture plates (Corning) containing 100micro-litre of StemSpan Serum-Free Expansion Medium (Stemcell technologies) supplemented with 100ng/ml of human SCF and FLT-3L, 50ng/ml of human TPO, 20ng/ml of human IL-3, IL-6 and G-CSF (all cytokines from Peprotech) and 50U/ml of penicillin and 50μg/ml of streptomycin (Sigma). Cells were incubated at 37 degrees C in a humidified atmosphere with 5% CO2 in air. After 5 days in culture, another 100micro litres of cytokine-containing medium were added. 13 days after seeding, clones B6 and G2 had expanded to approx. 105 cells and were selected for whole genome sequencing (2x101bp, paired-end, Illumina HiSeq2500) after tagmentation-based library preparation (see Extended Experimental Procedures) for clone B6 and standard library preparation for clone G2. For germline-control ~106 unsorted bone marrow mononuclear cells from the same donor were used for sequencing. An average of 30-fold sequence coverage for each the clones and the matching control were obtained.L4clone: A progenitor cell clone was raised from a peripheral blood sample of the 39 year old healthy female. Frozen peripheral blood mononuclear cells (PBMCs) were isolated from 2 ml heparinised peripheral blood via Ficoll Paque density centrifugation. A methylcellulose assay was performed as described earlier (Weisse et al., 2012). In brief, non-adherent mononuclear cells were incubated in the presence of the recombinant human cytokines IL-3, IL-5 and GM-CSF (R&D systems) over 14 days to induce colony formation. Colonies were detected under an inverted light microscope, and plucked by a pipette when colonies had approximately 10,000 cells/CFU. Each colony was washed three times in PBS and finally frozen as a cell pellet in -80 degrees C. Genomic DNA was isolated using the QIAamp DNA micro kit according to the instructions of the manufacturer (Qiagen, Hilden, Germany). Whole genome sequencing (2x101bp, paired-end, Illumina HiSeq2500) was performed for colony 4 after tagmentation-based library preparation and resulted in 15-fold sequence coverage for each the colony and the matching whole blood.

Dataset EGAD00001000666

ProHealth: Kaiser Permanente Genome-wide Association Study of Prostate Cancer

A genome-wide association study (GWAS) of prostate cancer (PCa) was conducted in Kaiser Permanente (KP) Northern California health plan members (7,783 cases, 38,595 controls; 80.3% non-Hispanic white, 4.9% African-American, 7.0% East Asian, and 7.8% Latino) [PMID: 26034056]. The data for these members were drawn from three KP cohort studies: Research Program in Genes, Environment and Health (RPGEH) ProHealth, and California Men's Health Study (CMHS) (described further under Study History). Four custom arrays were designed for genotyping, one for each of the four major race-ethnicity groups in the RPGEH cohort: African Americans, East Asians, Latinos, and Non-Hispanic Whites. The number of SNPs and SNP content varied by array, with SNP content designed to maximize the genome-wide coverage of low frequency and more common variants specific to the different race-ethnicity groups, including newly identified SNPs from sequencing projects, and SNPs with established associations with disease phenotypes and risk factors [PMIDs: 21565264, 21903159]. Within the total study cohort, n=34,736 completed a consent which permitted deposition of data to NIH. Genotyping followed the same general procedure described in [PMIDs: 26092718, plus additional quality control (QC) steps for the additional men, in order to control for potential batch and kit effects, described in [PMID: 26034056. Briefly, we first repeated the filters described in [PMID: 26092718] for all four arrays (EUR, LAT, EAS, AFR). Then, on an array-wise basis, we removed SNPs with MAF<0.01, with a call rate<95%, or with Hardy-Weinberg Equilibrium (HWE) p-value in homogeneous groups<1x10ˆ-5. Furthermore, on the EUR array, to adjust for potential kit effect, we conducted a GWAS of kit, and removed those kit associated SNPs with p<1x10ˆ-6; we also re-genotyped each of the new samples (those not genotyped with the original GERA data) with some of the original GERA data, and removed SNPs with >13/1,268 (1%) mismatches. For the AFR array, to adjust potential plate batch issues, we conducted a GWAS of whether an individual was in the original GERA data vs. in the newly genotyped data and removed those batch-associated SNPs with p<0.05 (we used a stronger threshold than that used for the EUR array because there were fewer individuals on the AFR array); we also re-genotyped each of the new samples with the original GERA data and removed SNPs with >2/78 (2.6%). After the QC described above, imputation was performed as described in [PMID: 26034056]. Imputation was performed on an array-wise basis, pre-phasing with SHAPE-IT v2.5 [PMID: 22138821], and imputing from the 1000 Genomes Project October 2014 release as a cosmopolitan reference panel with IMPUTE2 [PMID: 22384356]. In addition to the GWAS described above, a nested exome-wide association study (EWAS) of PCa was also conducted (7,489 cases, 7,323 controls; 78% non-Hispanic white, 9% African-American, 3% East Asian, 6% Latino, 4% Other). A custom EWAS array primarily focused on rare variants was designed for genotyping that complemented the GWAS arrays [PMID: 26034056]. The EWAS array content included missense and loss-of-function mutations, and rare exonic mutations from The Cancer Genome Atlas (TCGA) and dbGaP prostate cancer tumor exomes [PMID: 26544944; PMID: 26544944]. Much of the EWAS array design content overlapped with the probesets on the UK Biobank Affymetrix Axiom array [PMID: 30305743]. Genotyping and QC steps taken to filter out samples exhibiting low quality and variants with low call rates are described in Emami et al., 2020 [biorXiv]. The resulting EWAS array genotypes are provided here.

Study phs001221

The Collaborative Study on the Genetics of Alcoholism (COGA)

COGA is a family study of alcoholism, in which the subjects have been drawn from the Collaborative Study on the Genetics of Alcoholism (COGA), a large, ongoing family-based study that includes subjects from seven sites around the US. COGA has gathered detailed, standardized data on study participants, including diagnostic and neurophysiological assessments. This project has already proved successful in identifying several genes that influence the risk for alcoholism and neurophysiological endophenotypes, which have been independently replicated. COGA data were included as part of two Genetic Analysis Workshops, and the phenotypes are familiar to the genetics community. Alcoholic probands were recruited from treatment facilities, assessed by personal interview, and after securing permission, other family members were also assessed. A set of comparison families was drawn from the same communities as the families recruited through an alcoholic proband. Assessment involved a detailed personal interview developed for this project, the Semi-Structured Assessment for the Genetics of Alcoholism (SSAGA), which gathers detailed information on alcoholism related symptoms along with other drugs and psychiatric symptoms. Many participants also came to the laboratories for electroencephalographic studies. Neurophysiological features that have been shown to be useful endophenotypes for which we have linkage and in some cases association results, are included for a subset of the case-control sample: the beta power of the resting electroencephalogram (EEG), the P3(00) amplitude of the visual event-related potential (ERP), and the theta and delta event-related oscillations (EROs) underlying the P3. As part of COGA, a set of informative families was selected to have Genome-Wide Association data obtained within families. Genotyping was performed using the Illumina Human OmniExpress array 12.VI to genotype 2,282 subjects selected from 118 densely affected families. Genotyping was performed at the Genome Technology Access Center at Washington University School of Medicine in St. Louis. In addition, we also included genotypes for subjects (n=275 subjects) from these 118 families who were genotyped in a previous case-control GWAS using the Illumina 1M array. For quality control purposes, 51 of the 275 subjects were genotyped again on the Illumina Human OmniExpress array at the Washington University School of Medicine core facility.In addition, exome sequencing data on a subset of individuals with GWAS were added in version 2 (v2). For v2, a subset had 30X Whole Genome Sequencing (WGS) as part of the NIDA Sequencing Initiative. The subset contained two distinct sets: Sibling pairs where one sibling had at least two dependence diagnoses in the set (alcohol, cannabis, cocaine, and opioid), and the other had none, and non-related Case-Control pairs matched for age and ethnicity where the cases had alcohol and at least 2 other dependence diagnoses and controls had none. After sequencing, some sibling pairs are re-classified as half siblings. Three VCF files (small variants, structural variants, and copy number variations) are provided. Additional substance use variables are made available in v2. We note that the full sample data are deposited in four dbGaP submissions and the sequenced samples are split across all four: CIDR: Collaborative Study on the Genetics of Alcoholism Case Control Study [phs000125]. GWAS data on cases (primarily probands) and controls drawn from the families. Families with highest density of alcohol dependence and/or extreme event-related oscillation data [phs000763]. GWAS data on 119 extended families of European descent are available here, along with extensive documentation. Study on the Genetics of Alcoholism (COGA): African American Family GWAS [phs000976]. GWAS data on all available COGA families of African descent are available. COGA: Smokescreen GWAS [phs001208]. GWAS data on all remaining COGA DNA samples, primarily of other racial background, were genotyped on the Smoke Screen array. A listing of all sequenced pairs is provided in the documentation to facilitate the merging of these samples.

Study phs000763

University of Washington Cerebrospinal Fluid Biomarker Study for Parkinson disease

There is a clear need to develop biomarkers for Parkinson disease (PD) diagnosis and monitoring disease progression. In this study we evaluated cerebrospinal fluid (CSF) proteins, which are known to be critically involved in PD or identified in our preliminary profiling studies, aptamers, and RNAs as potential PD biomarkers. Access to subjects for this study was via the Pacific Northwest Udall Center (PANUC) and the Alzheimer's Disease Research Center (ADRC) at the University of Washington and Oregon Health and Sciences University (OHSU). Using CSF samples from 30 well-characterized patients with PD and 30 age-, sex-matched healthy controls, we prepared RNA seq libraries and performed deep sequencing of all RNA species, including small and long RNA, mRNAs, noncoding RNAs and differentially spliced transcripts. We then tried several methods for RNAseq data analysis to optimize our analysis pipeline. We identified a total of 3381 transcripts corresponding to 182 long intergenic RNAs (LincRNAs), 11 microRNAs (miRNAs), 2861 protein-coding transcripts, 200 pseudogenes and 127 antisense RNAs; some of them were differentially expressed between PD and control groups. Selected differentially expressed RNAs have been validated in the same set of CSF samples using real-time PCR (RT-PCR). Further validations in independent, larger cohorts of samples are still ongoing. Our results obtained so far suggested that CSF proteins and RNAs could be used as good indexes for PD diagnosis and disease severity/progression. This study is a part of the NIDDS-funded Parkinson's Disease Biomarkers Program (PDBP).

Study phs000901

eMERGE: Genetics of Complex Pediatric Disorders from the Center for Applied Genomics

The Center for Applied Genomics (CAG) is a specialized Center of Emphasis at the Children's Hospital of Philadelphia (CHOP) with the primary goal of translating basic research findings to medical innovations. The mission of CAG is to develop new and better ways to diagnose and treat children affected by rare and complex medical disorders. We aim to discover genetic causes for the most prevalent diseases of childhood including asthma, autism, diabetes, epilepsy, obesity, schizophrenia, pediatric cancer, and a range of rare diseases. Ultimately, our objective is to generate new diagnostic tests and to guide physicians to the most appropriate therapies. The CAG is one of the world's largest genetics research programs in pediatrics, and the only center at a pediatric hospital to have established a large-scale biobank of genotyped samples. The electronic Medical Records and Genomics (eMERGE) Network is a consortium of 9 clinical sites with EMR linked DNA biobanks, including Northwestern University and its NUgene biobank, funded by the NHGRI (National Human Genome Research Institute) to investigate the use of electronic medical record systems for genomic research. The goal of eMERGE is to conduct genome-wide association studies in approximately 100,000 individuals using EMR-derived phenotypes and DNA from linked biorepositories. Using electronic phenotyping methods, the consortium has been and is using DNA samples from all participating sites to explore the genetic determinants of approximately 80 phenotypes, including both diseases and traits, for which the electronic phenotyping algorithms have or are being published on PheKB.org.

Study phs001165

Search for new loci and low-frequency variants influencing glioma risk by exome-array analysis

To identify protein altering variants (PAVs) for glioma we analysed Illumina HumanExome BeadChip exome array data on 1,882 glioma cases and 8,079 controls from three independent European populations. In addition to single variant tests we incorporated information on the predicted functional consequences of PAVs and analysed sets of genes with a higher likelihood of having a role in glioma on the basis of the profile of somatic mutations documented by large-scale sequencing initiatives. Globally tThere was a strong relationship between effect size and SNPs predicted to be damaging (P=2.29x10-49); however, these variants which are most likely to impact on risk, are rare (MAF<5%). While no single variant showed an association which was statistically significant at the genomewide threshold a number of represented promising Notable associations - were seen with the BRCA2:c.9976A>T, p.(Lys3326Ter)BRCA2 p.Lys332X variant, which has been shown documented to influence breast and lung cancer risks (OR=2.3, P=4.00x10-4 for glioblastoma [GBM]) and IDH2:c.782G>A, p.(Arg261His)IDH2 p.Arg261His (OR=3.21, P=7.67x10-3, for non-GBM). Additionally, gene burden tests revealed a statistically significant association for HARS2 and risk of GBM (P=2.20x10-6). Genome scans of low frequency PAVs represent a complementary strategy to identify disease-causing variants compared with scans based on tagSNPs. Strategies to lessen the multiple testing burden by restricting analysis to PAVs with higher priors affords an opportunity to maximise study power.

Study EGAS00001001258

Versatile workflow for cell-type resolved transcriptional and epigenetic profiles from cryopreserved human lung

The complexity of the lung microenvironment and changes in cellular composition during disease progression make it exceptionally hard to understand molecular mechanisms leading to the development of chronic lung diseases. Although recent advances in cell-type resolved and single-cell sequencing approaches hold great promise for studying complex diseases, their implementation relies on local access to fresh tissue, as traditional tissue storage methods do not allow viable cell isolation. To overcome these hurdles, we developed a novel, versatile workflow that allows long-term storage of human lung tissue with high cell viability, permits thorough sample quality check before cell isolation, and is compatible with sequencing-based profiling, including single-cell approaches. We demonstrate that cryopreservation is suitable for the isolation of multiple cell types from human lung and is applicable to both healthy and diseased tissue, including COPD and tumour samples. Basal cells isolated from cryopreserved airways retain the ability to differentiate, indicating that cellular identity is not altered by cryopreservation. Importantly, using RNA sequencing and EPIC Array, we show that genome-wide gene expression and DNA methylation signatures are preserved upon cryopreservation, emphasizing the suitability of our workflow for -omics profiling of human lung cells. In addition, we obtained high-quality single-cell RNA sequencing data of cells isolated from the cryopreserved human lung, demonstrating that cryopreservation empowers single-cell approaches. Overall, thanks to its simplicity, our cryopreservation workflow is well-suited for prospective tissue collection by academic collaborators and biobanks, opening worldwide access to human tissue.

Study EGAS00001004477

Dental Caries: Whole Genome Association and Gene x Environment Studies

Dental caries (also known as tooth decay) remains the most common chronic disease of childhood, five times more common than asthma and seven times more common than environmental allergies, with more than 40% of children exhibiting caries when they enter kindergarten. In 2005, it was estimated that dental health care costs were approximately $84 billion, of which 60% or about $50 billion were related to treatment of dental caries. Although overall caries prevalence has declined over the last 40 years, dental caries in the primary dentition and mean caries rates in children ages 2-11 has increased markedly over the past 12 years. Childhood caries is a serious public health issue because of associated health problems and because disparities in oral health have led to substantially higher average disease prevalence among children in poverty and in under-served racial and ethnic groups. These issues are of such concern that in 2005, the American Academy of Pediatrics made children's oral health one of their top areas of focus, as it is for the majority of the NIDCR "Disparities Centers". The etiology of dental caries has been studied for many years. Multiple factors contribute to a person's risk for caries, including: 1) environmental factors such as diet, oral hygiene, fluoride exposure and the level of colonization of cariogenic bacteria and 2) host factors such as salivary flow, salivary buffering capacity, position of teeth relative to each other, surface characteristics of tooth enamel and depth of occlusal fissures on posterior teeth. In spite of all that is known about this disease, there are still individuals who appear to be more susceptible to caries and those who are extremely resistant, regardless of the environmental risk factors to which they are exposed, implying that genetic factors also play an important role in caries etiology. This conclusion is supported by studies in both humans and animals, with the most compelling evidence coming from studies of twins reared apart in which investigators found significant resemblance within monozygotic (MZ) but not dizygotic (DZ) twin pairs for percentage of teeth and surfaces restored or carious and estimated the genetic contribution to caries as 40%. Other recent studies of twins reared together estimated the heritability for caries, adjusted for age and gender, as ranging from 45-64%. Despite the strong evidence of a genetic component to risk for dental caries, there have been only a few studies of candidate genes in caries, and no published genome-wide scans. A comprehensive genome wide search is the only approach that will allow us to identify those genetic regions likely to harbor genes increasing the risk for dental caries, and eventually to identify the etiologic genes and to explore the interaction of those genes with microbiological, dietary, fluoride, and behavioral factors that are known to be associated with caries risk and progression. Therefore, the goal of this study is to perform genome-wide association (GWA) studies of dental caries with a large panel of SNP's (610,000) in families and individuals ascertained through multiple US sites (University of Pittsburgh and University of Iowa). The v2 release of this study includes 96 additional individuals who were genotyped with the CCDG: Dental Caries and CL/P in Guatemala project (dbGaP accession number phs000440) to augment the data initially presented here. These subjects were genotyped on the Illumina 610 platform to make their data comparable. This study is part of the Gene Environment Association Studies initiative (GENEVA, http://www.genevastudy.org), which was developed through the trans-NIH Genes, Environment, and Health Initiative (GEI). The overarching goal is to identify novel genetic factors that contribute to dental caries through large-scale genome-wide association studies of well-characterized families and individuals at multiple sites in the U.S. Genotyping was performed at the Johns Hopkins University Center for Inherited Disease Research (CIDR). The study was supported by the National Institute of Dental and Craniofacial Research (NIDCR, U01-DE018903). Data cleaning and harmonization were done at the GEI-funded GENEVA Coordinating Center at the University of Washington. Samples of some participants in this study were selected for further genomic analysis. These results will be available in the dbGaP CIDR study: Center for Oral Health Research in Appalachia (COHRA) Genomic Studies of Oral Health and Disease (phs001591).

Study phs000095

Risk Assessment of Cerebrovascular Events (RACE) Study

This study includes 1,220 cases with young onset stroke (stroke before age 60 years) who are participants of the larger RACE study. Risk Assessment of Cerebrovascular Events (RACE) is an on-going existing case-control study of stroke now involving over 5000 imaging confirmed cases of stroke and 5000 controls, recruited from seven centers in Pakistan. The study is aimed to investigate the genetic, biomarker and lifestyle determinants of stroke and its subtypes. Cases are eligible for inclusion in the study if they: (i) are aged at least 18 years; (ii) present with a sudden onset of neurological deficit respecting a vascular territory with sustained deficit at 24 hours verified by medical attention within 72 hours after onset (onset is defined by when the patient was last seen normal and not when found with deficit); and (iii) the diagnosis is supported by CT/MRI; and (iv) present with a Modified Rankin Score < 2 prior to the stroke. Findings from patient's history, 12-lead ECG and CT or MRI of the brain. The mandatory procedures for inclusion in this investigation are: (i) clinical verification of cerebrovascular event within 72 hours of onset; (ii) neuroimaging CT (non-contrast) or MRI (MRI is not a mandatory investigation but recorded whenever ordered by the attending physician); and (iii) 12-lead ECG. All other ancillary investigations ordered by the attending physician are recorded as well. The TOAST classification method is used to classify ischemic stroke based on aetiology whereas the Oxfordshire classification is used to classify stroke neuro-anatomically. Control participants for this subset of young onset stroke were individuals enrolled in the Pakistan Risk of Myocardial Infarction Study (PROMIS), a case-control study of acute MI based in Pakistan. RACE capitalizes on the genetic data (including information on GWAS) that has already been collected from the healthy participants enrolled in PROMIS. RACE and PROMIS share similar methodology of recruitment. Participants from both these investigations are derived from similar catchment areas, hence providing an attractive opportunity for RACE to utilize PROMIS controls as common controls for genetic investigations. Controls in PROMIS were recruited following procedures and inclusion criteria as adopted for RACE cases. In order to minimize any potential selection biases, PROMIS controls selected for this stroke substudy were frequency matched to RACE cases based on age and gender and were recruited in the following order of priority: (1) non-blood related or blood related visitors of patients of the out-patient department; (2) non-blood related visitors of stroke patients; (3) patients of the out-patient department presenting with minor complaints (e.g. back pain, minor gastric complaints). Control subjects from the PROMIS study were genotyped at the Wellcome Trust Sanger Institute on the Illumina 660W Quad array. The Center for Non-Communicable Diseases, Pakistan, serves as the coordinating center for both RACE and PROMIS. More information on these research investigations can be found at www.cncdpk.com. This young onset stroke component to the RACE study was funded through the Gene Environment Association Studies initiative (GENEVA, www.genevastudy.org as one of three studies designed to assess the genetics of young onset stroke and modification of genetic effects by smoking. GENEVA is part of the trans-NIH Genes, Environment, and Health Initiative (GEI). Genotyping of 1,220 young onset stroke cases was performed at the Johns Hopkins University Center for Inherited Disease Research (CIDR). Data cleaning and harmonization were done at the GEI-funded GENEVA Coordinating Center at the University of Washington. This study is part of the Gene Environment Association Studies initiative (GENEVA, http://www.genevastudy.org) funded by the trans-NIH Genes, Environment, and Health Initiative (GEI). The overarching goal is to identify novel genetic factors that contribute to stroke through large-scale genome-wide association studies of cases and controls recruited within Pakistan. Genotyping was performed at the Johns Hopkins University Center for Inherited Disease Research (CIDR). Data cleaning and harmonization were done at the GEI-funded GENEVA Coordinating Center at the University of Washington.

Study phs000456

CentralAfricanCMC_Pemberton

The CentralAfricanCMC_Pemberton dataset encompasses 153,798 SNPs from the Illumina Cardio-MetaboChip (Voight et al. 2012) genotyped in 406 individuals from 19 Central African Populations from Gabon, Cameroon, Centralafrican Republic and Uganda). Individual phenotypic and cultural information at the individual level for this data set encompass gender, lifestyle (hunter-gatherer or farmer), and, when available, stature phenotype (standing height in cm, sitting-height in cm, and weight in kg). Other cultural, linguistic, and geographical location information about the sampled populations can be found in Pemberton et al. , Human Genetics, 2018 (https://doi.org/10.1007/s00439-018-1902-3).This dataset can only be accessed and used for non-commercial research purposes with a finality complying with the informed consent provided by Central African donors for the study of human evolutionary history only.

Dataset EGAD00010001584

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