scholarly journals Using synthetic chromosome controls to evaluate the sequencing of difficult regions within the human genome

2022 ◽  
Vol 23 (1) ◽  
Author(s):  
Andre L. M. Reis ◽  
Ira W. Deveson ◽  
Bindu Swapna Madala ◽  
Ted Wong ◽  
Chris Barker ◽  
...  
Keyword(s):  
Human Genome ◽  
Comprehensive Evaluation ◽  
Diagnostic Yield ◽  
Ground Truth ◽  
Low Complexity ◽  
Systematic Bias ◽  
Preparation Methods ◽  
Synthetic Dna ◽  
Bioinformatic Tools ◽  
Sequencing Technologies

Abstract Background Next-generation sequencing (NGS) can identify mutations in the human genome that cause disease and has been widely adopted in clinical diagnosis. However, the human genome contains many polymorphic, low-complexity, and repetitive regions that are difficult to sequence and analyze. Despite their difficulty, these regions include many clinically important sequences that can inform the treatment of human diseases and improve the diagnostic yield of NGS. Results To evaluate the accuracy by which these difficult regions are analyzed with NGS, we built an in silico decoy chromosome, along with corresponding synthetic DNA reference controls, that encode difficult and clinically important human genome regions, including repeats, microsatellites, HLA genes, and immune receptors. These controls provide a known ground-truth reference against which to measure the performance of diverse sequencing technologies, reagents, and bioinformatic tools. Using this approach, we provide a comprehensive evaluation of short- and long-read sequencing instruments, library preparation methods, and software tools and identify the errors and systematic bias that confound our resolution of these remaining difficult regions. Conclusions This study provides an analytical validation of diagnosis using NGS in difficult regions of the human genome and highlights the challenges that remain to resolve these difficult regions.

scholarly journals One in seven pathogenic variants can be challenging to detect by NGS: an analysis of 450,000 patients with implications for clinical sensitivity and genetic test implementation

2021 ◽  
Author(s):  
Stephen E. Lincoln ◽  
Tina Hambuch ◽  
Justin M. Zook ◽  
Sara L. Bristow ◽  
Kathryn Hatchell ◽  
...  
Keyword(s):  
Diagnostic Yield ◽  
Copy Number Variants ◽  
Low Complexity ◽  
Clinical Genetic ◽  
Clinical Sensitivity ◽  
Pathogenic Variants ◽  
Wide Range ◽  
Large Indels ◽  
Next Generation Sequencing Ngs ◽  
The Impact

Abstract Purpose To evaluate the impact of technically challenging variants on the implementation, validation, and diagnostic yield of commonly used clinical genetic tests. Such variants include large indels, small copy-number variants (CNVs), complex alterations, and variants in low-complexity or segmentally duplicated regions. Methods An interlaboratory pilot study used synthetic specimens to assess detection of challenging variant types by various next-generation sequencing (NGS)–based workflows. One well-performing workflow was further validated and used in clinician-ordered testing of more than 450,000 patients. Results In the interlaboratory study, only 2 of 13 challenging variants were detected by all 10 workflows, and just 3 workflows detected all 13. Limitations were also observed among 11 less-challenging indels. In clinical testing, 21.6% of patients carried one or more pathogenic variants, of which 13.8% (17,561) were classified as technically challenging. These variants were of diverse types, affecting 556 of 1,217 genes across hereditary cancer, cardiovascular, neurological, pediatric, reproductive carrier screening, and other indicated tests. Conclusion The analytic and clinical sensitivity of NGS workflows can vary considerably, particularly for prevalent, technically challenging variants. This can have important implications for the design and validation of tests (by laboratories) and the selection of tests (by clinicians) for a wide range of clinical indications.

scholarly journals DNA methylation marks inter-nucleosome linker regions throughout the human genome

2013 ◽  
Author(s):  
Benjamin P. Berman ◽  
Yaping Liu ◽  
Theresa K. Kelly
Keyword(s):  
Dna Methylation ◽  
Human Genome ◽  
Ctcf Binding ◽  
Gene Promoters ◽  
Cpg Dinucleotides ◽  
Sequencing Technologies ◽  
Nucleosome Organization ◽  
Genome Wide ◽  
A Genome ◽  
Sequencing Studies

Background: Nucleosome organization and DNA methylation are two mechanisms that are important for proper control of mammalian transcription, as well as epigenetic dysregulation associated with cancer. Whole-genome DNA methylation sequencing studies have found that methylation levels in the human genome show periodicities of approximately 190 bp, suggesting a genome-wide relationship between the two marks. A recent report (Chodavarapu et al., 2010) attributed this to higher methylation levels of DNA within nucleosomes. Here, we analyzed a number of published datasets and found a more compelling alternative explanation, namely that methylation levels are highest in linker regions between nucleosomes. Results: Reanalyzing the data from (Chodavarapu et al., 2010), we found that nucleosome-associated methylation could be strongly confounded by known sequence-related biases of the next-generation sequencing technologies. By accounting for these biases and using an unrelated nucleosome profiling technology, NOMe-seq, we found that genome-wide methylation was actually highest within linker regions occurring between nucleosomes in multi-nucleosome arrays. This effect was consistent among several methylation datasets generated independently using two unrelated methylation assays. Linker-associated methylation was most prominent within long Partially Methylated Domains (PMDs) and the positioned nucleosomes that flank CTCF binding sites. CTCF adjacent nucleosomes retained the correct positioning in regions completely devoid of CpG dinucleotides, suggesting that DNA methylation is not required for proper nucleosomes positioning. Conclusions: The biological mechanisms responsible for DNA methylation patterns outside of gene promoters remain poorly understood. We identified a significant genome-wide relationship between nucleosome organization and DNA methylation, which can be used to more accurately analyze and understand the epigenetic changes that accompany cancer and other diseases.

scholarly journals Recent advances in functional genome analysis

F1000Research ◽  
2018 ◽  
Vol 7 ◽  
pp. 1968 ◽  
Author(s):  
Roderic Guigo ◽  
Michiel de Hoon
Keyword(s):  
Functional Genomics ◽  
Human Genome ◽  
Large Scale ◽  
Massively Parallel Sequencing ◽  
Genome Project ◽  
Biological Traits ◽  
Cellular Behavior ◽  
Sequencing Technologies ◽  
Recent Advances ◽  
The Human Genome Project

At the beginning of this century, the Human Genome Project produced the first drafts of the human genome sequence. Following this, large-scale functional genomics studies were initiated to understand the molecular basis underlying the translation of the instructions encoded in the genome into the biological traits of organisms. Instrumental in the ensuing revolution in functional genomics were the rapid advances in massively parallel sequencing technologies as well as the development of a wide diversity of protocols that make use of these technologies to understand cellular behavior at the molecular level. Here, we review recent advances in functional genomic methods, discuss some of their current capabilities and limitations, and briefly sketch future directions within the field.

scholarly journals Comparison of Otolith Readability and Reproducibility of Counts of Translucent Zones Using Different Otolith Preparation Methods for Four Endemic Labeobarbus Species in Lake Tana, Ethiopia

Water ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 1336 ◽  
Author(s):  
Gebremedhin ◽  
Bekaert ◽  
Getahun ◽  
Bruneel ◽  
Anteneh ◽  
...  
Keyword(s):  
Management Strategies ◽  
Tropical Fish ◽  
Percentage Error ◽  
Systematic Bias ◽  
Preparation Technique ◽  
Lake Tana ◽  
Preparation Methods ◽  
Average Percentage ◽  
Average Percentage Error ◽  
Preparation Techniques

The analysis of fish age data is vital for the successful conservation of fish. Attempts to develop optimal management strategies for effective conservation of the endemic Labeobarbus species are strongly affected by the lack of accurate age estimates. Although methodological studies are key to acquiring a good insight into the age of fishes, up to now, there have not been any studies comparing different methods for these species. Thus, this study aimed at determining the best method for the endemic Labeobarbus species. Samples were collected from May 2016 to April 2017. Asteriscus otoliths from 150 specimens each of L. intermedius, L. tsanensis, L. platydorsus, and L. megastoma were examined. Six methods were evaluated; however, only three methods resulted in readable images. The procedure in which whole otoliths were first submerged in water, and subsequently placed in glycerol to take the image (MO1), was generally best. Except for L. megastoma, this method produced the clearest image as both the coefficient of variation and average percentage error between readers were lowest. Furthermore, except for L. megastoma, MO1 had high otolith readability and no systematic bias. Therefore, we suggest that MO1 should be used as the standard otolith preparation technique for the first three species, while for L. megastoma, other preparation techniques should be evaluated. This study provides a reference for researchers from Africa, particularly Ethiopia, to develop a suitable otolith preparation method for the different tropical fish species.

Direct-to-Consumer Genetic Testing

2011 ◽  
pp. 51-84 ◽  
Author(s):  
Richard A. Stein
Keyword(s):  
Human Genome ◽  
Human Genome Project ◽  
Cost Effective ◽  
Helical Structure ◽  
Genome Project ◽  
Next Generation ◽  
Double Helical Structure ◽  
Sequencing Technologies ◽  
Human Genome Sequencing ◽  
The Human Genome Project

The 1953 discovery of the DNA double-helical structure by James Watson, Francis Crick, Maurice Wilkins, and Rosalind Franklin, represented one of the most significant advances in the biomedical world (Watson and Crick 1953; Maddox 2003). Almost half a century after this landmark event, in February 2001, the initial draft sequences of the human genome were published (Lander et al., 2001; Venter et al., 2001) and, in April 2003, the International Human Genome Sequencing Consortium reported the completion of the Human Genome Project, a massive international collaborative endeavor that started in 1990 and is thought to represent the most ambitious undertaking in the history of biology (Collins et al., 2003; Thangadurai, 2004; National Human Genome Research Institute). The Human Genome Project provided a plethora of genetic and genomic information that significantly changed our perspectives on biomedical and social sciences. The sequencing of the first human genome was a 13-year, 2.7-billion-dollar effort that relied on the automated Sanger (dideoxy or chain termination) method, which was developed in 1977, around the same time as the Maxam-Gilbert (chemical) sequencing, and subsequently became the most frequently used approach for several decades (Sanger et al., 1975; Maxam & Gilbert, 1977; Sanger et al., 1977). The new generations of DNA sequencing technologies, known as next-generation (second generation) and next-next-generation (third generation) sequencing, which started to be commercialized in 2005, enabled the cost-effective sequencing of large chromosomal regions during progressively shorter time frames, and opened the possibility for new applications, such as the sequencing of single-cell genomes (Service, 2006; Blow, 2008; Morozova and Marra, 2008; Metzker, 2010).

scholarly journals ECG Sensor Verification System with Mean-Interval Algorithm for Handling Sport Issue

2016 ◽  
Vol 2016 ◽  
pp. 1-12 ◽  
Author(s):  
Kuo-Kun Tseng ◽  
Fufu Zeng ◽  
W. H. Ip ◽  
C. H. Wu
Keyword(s):  
Flash Memory ◽  
Comprehensive Evaluation ◽  
Heat Rate ◽  
Low Complexity ◽  
Biometric Verification ◽  
Interval Algorithm ◽  
Interval Approach ◽  
Multiple State ◽  
Card System ◽  
New Framework

With the development of biometric verification, we proposed a new algorithm and personal mobile sensor card system for ECG verification. The proposed new mean-interval approach can identify the user quickly with high accuracy and consumes a small amount of flash memory in the microprocessor. The new framework of the mobile card system makes ECG verification become a feasible application to overcome the issues of a centralized database. For a fair and comprehensive evaluation, the experimental results have been tested on public MIT-BIH ECG databases and our circuit system; they confirm that the proposed scheme is able to provide excellent accuracy and low complexity. Moreover, we also proposed a multiple-state solution to handle the heat rate changes of sports problem. It should be the first to address the issue of sports in ECG verification.

The chicken model organism for epigenomic research

Genome ◽  
2020 ◽  
Author(s):  
Tasnim H. BEACON ◽  
James R DAVIE
Keyword(s):  
High Throughput Sequencing ◽  
Chicken Genome ◽  
Regulation Of Gene Expression ◽  
Model Organism ◽  
Bioinformatic Tools ◽  
Sequencing Technologies ◽  
Trace Back ◽  
Human Orthologs ◽  
Genomic Studies ◽  
Chicken Model

The chicken model organism has advanced the areas of developmental biology, virology, immunology, oncology, epigenetic regulation of gene expression, conservation biology, and genomics of domestication. Further, the chicken model organism has aided in our understanding of human disease. Through the recent advances in high-throughput sequencing and bioinformatic tools, researchers have successfully identified sequences in the chicken genome that have human orthologs, improving mammalian genome annotation. In this review, we highlight the importance of chicken as an animal model in basic and pre-clinical research. We will present the importance of chicken in poultry epigenetics and in genomic studies that trace back to their ancestor, the last link between human and chicken tree of life. There are still many genes of unknown function in the chicken genome yet to be characterized. By taking advantage of recent sequencing technologies, it is possible to gain further insight into the chicken epigenome.

scholarly journals Prediction of driver variants in the cancer genome via machine learning methodologies

2020 ◽  
Author(s):  
Mark F Rogers ◽  
Tom R Gaunt ◽  
Colin Campbell
Keyword(s):  
Machine Learning ◽  
Cell Proliferation ◽  
Human Genome ◽  
Human Cancer ◽  
Cancer Genome ◽  
Prediction Tools ◽  
Sequencing Technologies ◽  
Bioinformatics Tools

Abstract Sequencing technologies have led to the identification of many variants in the human genome which could act as disease-drivers. As a consequence, a variety of bioinformatics tools have been proposed for predicting which variants may drive disease, and which may be causatively neutral. After briefly reviewing generic tools, we focus on a subset of these methods specifically geared toward predicting which variants in the human cancer genome may act as enablers of unregulated cell proliferation. We consider the resultant view of the cancer genome indicated by these predictors and discuss ways in which these types of prediction tools may be progressed by further research.

scholarly journals Comprehensive Evaluation and Optimization of Amplicon Library Preparation Methods for High-Throughput Antibody Sequencing

PLoS ONE ◽  
2014 ◽  
Vol 9 (5) ◽  
pp. e96727 ◽  
Author(s):  
Ulrike Menzel ◽  
Victor Greiff ◽  
Tarik A. Khan ◽  
Ulrike Haessler ◽  
Ina Hellmann ◽  
...  
Keyword(s):  
High Throughput ◽  
Comprehensive Evaluation ◽  
Library Preparation ◽  
Preparation Methods ◽  
Amplicon Library

scholarly journals ‘There and back again’: revisiting the pathophysiological roles of human endogenous retroviruses in the post-genomic era

2013 ◽  
Vol 368 (1626) ◽  
pp. 20120504 ◽  
Author(s):  
Gkikas Magiorkinis ◽  
Robert Belshaw ◽  
Aris Katzourakis
Keyword(s):  
Human Genome ◽  
High Throughput Sequencing ◽  
Immune Escape ◽  
Genomic Sequence ◽  
Sequence Data ◽  
Endogenous Retroviruses ◽  
Human Endogenous Retroviruses ◽  
The Past ◽  
Sequencing Technologies ◽  
Research Questions

Almost 8% of the human genome comprises endogenous retroviruses (ERVs). While they have been shown to cause specific pathologies in animals, such as cancer, their association with disease in humans remains controversial. The limited evidence is partly due to the physical and bioethical restrictions surrounding the study of transposons in humans, coupled with the major experimental and bioinformatics challenges surrounding the association of ERVs with disease in general. Two biotechnological landmarks of the past decade provide us with unprecedented research artillery: (i) the ultra-fine sequencing of the human genome and (ii) the emergence of high-throughput sequencing technologies. Here, we critically assemble research about potential pathologies of ERVs in humans. We argue that the time is right to revisit the long-standing questions of human ERV pathogenesis within a robust and carefully structured framework that makes full use of genomic sequence data. We also pose two thought-provoking research questions on potential pathophysiological roles of ERVs with respect to immune escape and regulation.

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