scholarly journals Evaluation of Mayer-Rokitansky-Kuster-Hauser (MRKH) Patient Families by Whole Genome Sequencing

2021 ◽  
Vol 5 (Supplement_1) ◽  
pp. A501-A502
Author(s):  
Michael P Dougherty ◽  
Lynn P Chorich ◽  
Lawrence Clarke Layman
Keyword(s):  
Whole Genome Sequencing ◽  
Candidate Genes ◽  
Genome Sequencing ◽  
De Novo ◽  
Splice Variants ◽  
Family Members ◽  
Gene Variants ◽  
Whole Genome ◽  
Missense Variants ◽  
Coding Regions

Abstract Introduction: MRKH is a characterized by the congenital absence of the uterus and vagina in 46,XX individuals. A subset of these patients also has associated renal, skeletal, cardiac and/or auditory defects. Familial cases suggest a genetic component, but to date only pathogenic variants in WNT4 and HNF1B have been confirmed. We hypothesize that de novo heterozygous variants in candidate genes will be present in some patients with MRKH. Methods: DNAs from 30 quads (an MRKH proband and three relatives) were subjected to whole genome sequencing (WGS), and heterozygous variants in coding regions with < 0.02 frequency were filtered by two different methods. In the first approach, variants were filtered by 1) top consequence variant (splice site, stop-gain, frameshift, and missense, respectively); 2) impact score; 3) mapping quality; 4) cytobands; 5) intolerance; 6) de novo variants; and 7) plausibility based on familial genotype. The second approach considered only heterozygous variants found in the proband and absent in all other family members, which were then filtered by top consequence (splice donor and acceptor sites, stop-gain, frameshift). Results: Five pedigrees were excluded for inadequate sequence in one or more individuals. 55,033 variants in coding regions with < 2% frequency were identified in the 25 remaining quads for analysis. Using the first approach, 42 candidate gene variants in 32 genes were identified - 12 splice variants, 10 stop-gains, 15 frameshift variants and 5 missense variants. Of these, MUC22 contained 2 missense variants from different families. Additionally, DICER1 had multiple splice variants and is essential for mouse urogenital tract development. In the second approach, 39 candidate genes were identified—6 splice variants in 6 genes, 18 stop-gains in 17 genes, and 17 frameshift variants in 16 genes. Zinc finger genes (ZNF418, ZNF646, ZNF135, and ZNF772) comprised the most frequent class of the 39 genes. Two genes (MIR4436A and ZNF418) contained attractive variants in two different families. Conclusion: WGS has been shown to improve detection of gene variants in coding regions, more so than whole exome sequencing (WES). We previously performed WES on 111 MRKH probands without family members and analyzed variants in candidate genes suggested by mouse and preliminary human studies. Interestingly, in this study, only three genes overlapped with previously suspected candidate genes. Here, we identified new candidates based upon potential deleteriousness. These candidate genes will be studied further in our families to determine their role in Mullerian development.

scholarly journals A statistical framework for mapping risk genes from de novo mutations in whole-genome sequencing studies

2016 ◽  
Author(s):  
Yuwen Liu ◽  
Yanyu Liang ◽  
A. Ercument Cicek ◽  
Zhongshan Li ◽  
Jinchen Li ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
De Novo ◽  
Genome Wide Association Studies ◽  
Whole Genome ◽  
Major Advance ◽  
Risk Genes ◽  
Coding Sequences ◽  
Coding Regions ◽  
Statistical Framework

AbstractAnalysis of de novo mutations (DNMs) from sequencing data of nuclear families has identified risk genes for many complex diseases, including multiple neurodevelopmental and psychiatric disorders. Most of these efforts have focused on mutations in protein-coding sequences. Evidence from genome-wide association studies (GWAS) strongly suggests that variants important to human diseases often lie in non-coding regions. Extending DNM-based approaches to non-coding sequences is, however, challenging because the functional significance of non-coding mutations is difficult to predict. We propose a new statistical framework for analyzing DNMs from whole-genome sequencing (WGS) data. This method, TADA-Annotations (TADA-A), is a major advance of the TADA method we developed earlier for DNM analysis in coding regions. TADA-A is able to incorporate many functional annotations such as conservation and enhancer marks, learn from data which annotations are informative of pathogenic mutations and combine both coding and non-coding mutations at the gene level to detect risk genes. It also supports meta-analysis of multiple DNM studies, while adjusting for study-specific technical effects. We applied TADA-A to WGS data of ∼300 autism family trios across five studies, and discovered several new autism risk genes. The software is freely available for all research uses.

scholarly journals Whole genome sequencing identifies variants associated with sarcoidosis in a family with a high prevalence of sarcoidosis

2021 ◽  
Author(s):  
Daan Fritz ◽  
Bart Ferwerda ◽  
Matthijs C. Brouwer ◽  
Diederik van de Beek
Keyword(s):  
Whole Genome Sequencing ◽  
Candidate Genes ◽  
Genome Sequencing ◽  
Family Members ◽  
Index Patient ◽  
High Rate ◽  
Whole Genome ◽  
Loss Of Function ◽  
Treatment Resistant ◽  
High Prevalence

Abstract Objective We studied genetic risk factors associated with sarcoidosis within a family with a high prevalence of this disease. Methods We studied 41 members of a family with a high rate of sarcoidosis, including an index patient with treatment-resistant neurosarcoidosis. Whole genome sequencing was performed for six affected family members and variations associated with loss of function were filtered out as candidate genes. Findings were validated by using amplicon sequencing within all 41 family members with DNA available and candidate genes were screened on absence and presence within the sarcoidosis affected and non-affected. Results Family members (n = 61) from 5 generations were available for participation including 13 subjects diagnosed with sarcoidosis (20%). Analyses identified 36 candidate variants within 34 candidate genes. Variations within three of these genes (JAK2, BACH2, and NCF1) previously have been associated with autoimmune diseases. Conclusions We identified 34 genes with a possible role in the etiology of sarcoidosis, including JAK2. Our results may suggest evaluation of JAK inhibitors in treatment-resistant sarcoidosis. Key Points• JAK2 has a potential role in the etiology of sarcoidosis and is a potential therapeutic target.• We identified 33 additional candidate genes of which BACH2 and NCF1 have been previously associated with autoimmune disease.

scholarly journals Effective variant filtering and expected candidate variant yield in studies of rare human disease

2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Brent S. Pedersen ◽  
Joe M. Brown ◽  
Harriet Dashnow ◽  
Amelia D. Wallace ◽  
Matt Velinder ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Rare Disease ◽  
Genome Sequencing ◽  
Autosomal Dominant ◽  
De Novo ◽  
Autosomal Dominant Inheritance ◽  
Compound Heterozygous ◽  
Whole Genome ◽  
Dominant Inheritance ◽  
Family Based

AbstractIn studies of families with rare disease, it is common to screen for de novo mutations, as well as recessive or dominant variants that explain the phenotype. However, the filtering strategies and software used to prioritize high-confidence variants vary from study to study. In an effort to establish recommendations for rare disease research, we explore effective guidelines for variant (SNP and INDEL) filtering and report the expected number of candidates for de novo dominant, recessive, and autosomal dominant modes of inheritance. We derived these guidelines using two large family-based cohorts that underwent whole-genome sequencing, as well as two family cohorts with whole-exome sequencing. The filters are applied to common attributes, including genotype-quality, sequencing depth, allele balance, and population allele frequency. The resulting guidelines yield ~10 candidate SNP and INDEL variants per exome, and 18 per genome for recessive and de novo dominant modes of inheritance, with substantially more candidates for autosomal dominant inheritance. For family-based, whole-genome sequencing studies, this number includes an average of three de novo, ten compound heterozygous, one autosomal recessive, four X-linked variants, and roughly 100 candidate variants following autosomal dominant inheritance. The slivar software we developed to establish and rapidly apply these filters to VCF files is available at https://github.com/brentp/slivar under an MIT license, and includes documentation and recommendations for best practices for rare disease analysis.

scholarly journals Evaluating coverage bias in next-generation sequencing of Escherichia coli

PLoS ONE ◽  
2021 ◽  
Vol 16 (6) ◽  
pp. e0253440
Author(s):  
Samantha Gunasekera ◽  
Sam Abraham ◽  
Marc Stegger ◽  
Stanley Pang ◽  
Penghao Wang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
De Novo ◽  
Fragment Size ◽  
Gc Content ◽  
Main Concern ◽  
Whole Genome ◽  
Assembly Quality ◽  
Coverage Bias

Whole-genome sequencing is essential to many facets of infectious disease research. However, technical limitations such as bias in coverage and tagmentation, and difficulties characterising genomic regions with extreme GC content have created significant obstacles in its use. Illumina has claimed that the recently released DNA Prep library preparation kit, formerly known as Nextera Flex, overcomes some of these limitations. This study aimed to assess bias in coverage, tagmentation, GC content, average fragment size distribution, and de novo assembly quality using both the Nextera XT and DNA Prep kits from Illumina. When performing whole-genome sequencing on Escherichia coli and where coverage bias is the main concern, the DNA Prep kit may provide higher quality results; though de novo assembly quality, tagmentation bias and GC content related bias are unlikely to improve. Based on these results, laboratories with existing workflows based on Nextera XT would see minor benefits in transitioning to the DNA Prep kit if they were primarily studying organisms with neutral GC content.

scholarly journals Deletion in the Cobalamin Synthetase W Domain–Containing Protein 1 Gene Is associated with Congenital Anomalies of the Kidney and Urinary Tract

2019 ◽  
Vol 31 (1) ◽  
pp. 139-147
Author(s):  
Shoichiro Kanda ◽  
Masaki Ohmuraya ◽  
Hiroyuki Akagawa ◽  
Shigeru Horita ◽  
Yasuhiro Yoshida ◽  
...  
Keyword(s):  
Urinary Tract ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Congenital Anomalies ◽  
Kidney Development ◽  
Family Members ◽  
Incomplete Penetrance ◽  
Whole Genome ◽  
Renal Hypoplasia ◽  
Unilateral Renal Agenesis

BackgroundResearchers have identified about 40 genes with mutations that result in the most common cause of CKD in children, congenital anomalies of the kidney and urinary tract (CAKUT), but approximately 85% of patients with CAKUT lack mutations in these genes. The anomalies that comprise CAKUT are clinically heterogenous, and thought to be caused by disturbances at different points in kidney development. However, identification of novel CAKUT-causing genes remains difficult because of their variable expressivity, incomplete penetrance, and heterogeneity.MethodsWe investigated two generations of a family that included two siblings with CAKUT. Although the parents and another child were healthy, the two affected siblings presented the same manifestations, unilateral renal agenesis and contralateral renal hypoplasia. To search for a novel causative gene of CAKUT, we performed whole-exome and whole-genome sequencing of DNA from the family members. We also generated two lines of genetically modified mice with a gene deletion present only in the affected siblings, and performed immunohistochemical and phenotypic analyses of these mice.ResultsWe found that the affected siblings, but not healthy family members, had a homozygous deletion in the Cobalamin Synthetase W Domain–Containing Protein 1 (CBWD1) gene. Whole-genome sequencing uncovered genomic breakpoints, which involved exon 1 of CBWD1, harboring the initiating codon. Immunohistochemical analysis revealed high expression of Cbwd1 in the nuclei of the ureteric bud cells in the developing kidneys. Cbwd1-deficient mice showed CAKUT phenotypes, including hydronephrosis, hydroureters, and duplicated ureters.ConclusionsThe identification of a deletion in CBWD1 gene in two siblings with CAKUT implies a role for CBWD1 in the etiology of some cases of CAKUT.

scholarly journals A combined RNA-seq and whole genome sequencing approach for identification of non-coding pathogenic variants in single families

2020 ◽  
Vol 29 (6) ◽  
pp. 967-979 ◽  
Author(s):  
Revital Bronstein ◽  
Elizabeth E Capowski ◽  
Sudeep Mehrotra ◽  
Alex D Jansen ◽  
Daniel Navarro-Gomez ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genetic Diagnosis ◽  
Genetic Diagnostics ◽  
Whole Genome ◽  
Unique Challenge ◽  
Coding Regions ◽  
Pathogenic Variants ◽  
Number Of Patients ◽  
Coding Variants

Abstract Inherited retinal degenerations (IRDs) are at the focus of current genetic therapeutic advancements. For a genetic treatment such as gene therapy to be successful, an accurate genetic diagnostic is required. Genetic diagnostics relies on the assessment of the probability that a given DNA variant is pathogenic. Non-coding variants present a unique challenge for such assessments as compared to coding variants. For one, non-coding variants are present at much higher number in the genome than coding variants. In addition, our understanding of the rules that govern the non-coding regions of the genome is less complete than our understanding of the coding regions. Methods that allow for both the identification of candidate non-coding pathogenic variants and their functional validation may help overcome these caveats allowing for a greater number of patients to benefit from advancements in genetic therapeutics. We present here an unbiased approach combining whole genome sequencing (WGS) with patient-induced pluripotent stem cell (iPSC)-derived retinal organoids (ROs) transcriptome analysis. With this approach, we identified and functionally validated a novel pathogenic non-coding variant in a small family with a previously unresolved genetic diagnosis.

scholarly journals Contributions of de novo variants to systemic lupus erythematosus

2020 ◽  
Vol 29 (1) ◽  
pp. 184-193 ◽  
Author(s):  
Jonas Carlsson Almlöf ◽  
Sara Nystedt ◽  
Aikaterini Mechtidou ◽  
Dag Leonard ◽  
Maija-Leena Eloranta ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Lupus Erythematosus ◽  
De Novo ◽  
Whole Genome ◽  
Gene Promoters ◽  
Single Nucleotide Variants ◽  
Systemic Lupus ◽  
Promoter Regions

AbstractBy performing whole-genome sequencing in a Swedish cohort of 71 parent-offspring trios, in which the child in each family is affected by systemic lupus erythematosus (SLE, OMIM 152700), we investigated the contribution of de novo variants to risk of SLE. We found de novo single nucleotide variants (SNVs) to be significantly enriched in gene promoters in SLE patients compared with healthy controls at a level corresponding to 26 de novo promoter SNVs more in each patient than expected. We identified 12 de novo SNVs in promoter regions of genes that have been previously implicated in SLE, or that have functions that could be of relevance to SLE. Furthermore, we detected three missense de novo SNVs, five de novo insertion-deletions, and three de novo structural variants with potential to affect the expression of genes that are relevant for SLE. Based on enrichment analysis, disease-affecting de novo SNVs are expected to occur in one-third of SLE patients. This study shows that de novo variants in promoters commonly contribute to the genetic risk of SLE. The fact that de novo SNVs in SLE were enriched to promoter regions highlights the importance of using whole-genome sequencing for identification of de novo variants.

scholarly journals A pilot study for channel catfish whole genome sequencing and de novo assembly

BMC Genomics ◽  
2011 ◽  
Vol 12 (1) ◽  
Author(s):  
Yanliang Jiang ◽  
Jianguo Lu ◽  
Eric Peatman ◽  
Huseyin Kucuktas ◽  
Shikai Liu ◽  
...  
Keyword(s):  
Pilot Study ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Channel Catfish ◽  
De Novo Assembly ◽  
De Novo ◽  
Whole Genome
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