scholarly journals Highly Sensitive Diagnosis of 43 Monogenic Forms of Diabetes or Obesity Through One-Step PCR-Based Enrichment in Combination With Next-Generation Sequencing

Diabetes Care ◽  
2013 ◽  
Vol 37 (2) ◽  
pp. 460-467 ◽  
Author(s):  
A. Bonnefond ◽  
J. Philippe ◽  
E. Durand ◽  
J. Muller ◽  
S. Saeed ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Next Generation ◽  
Highly Sensitive ◽  
One Step ◽  
Monogenic Forms Of Diabetes ◽  
Generation Sequencing

scholarly journals A customized scaffolds approach for the detection and phasing of complex variants by next-generation sequencing

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Qiandong Zeng ◽  
Natalia T. Leach ◽  
Zhaoqing Zhou ◽  
Hui Zhu ◽  
Jean A. Smith ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Variant Calling ◽  
Causative Mutation ◽  
Next Generation ◽  
Single Nucleotide ◽  
Highly Sensitive ◽  
Highly Sensitive Detection ◽  
Next Generation Sequencing Ngs ◽  
Reference Bias ◽  
Generation Sequencing

Abstract Next-generation sequencing (NGS) is widely used in genetic testing for the highly sensitive detection of single nucleotide changes and small insertions or deletions. However, detection and phasing of structural variants, especially in repetitive or homologous regions, can be problematic due to uneven read coverage or genome reference bias, resulting in false calls. To circumvent this challenge, a computational approach utilizing customized scaffolds as supplementary reference sequences for read alignment was developed, and its effectiveness demonstrated with two CBS gene variants: NM_000071.2:c.833T>C and NM_000071.2:c.[833T>C; 844_845ins68]. Variant c.833T>C is a known causative mutation for homocystinuria, but is not pathogenic when in cis with the insertion, c.844_845ins68, because of alternative splicing. Using simulated reads, the custom scaffolds method resolved all possible combinations with 100% accuracy and, based on > 60,000 clinical specimens, exceeded the performance of current approaches that only align reads to GRCh37/hg19 for the detection of c.833T>C alone or in cis with c.844_845ins68. Furthermore, analysis of two 1000 Genomes Project trios revealed that the c.[833T>C; 844_845ins68] complex variant had previously been undetected in these datasets, likely due to the alignment method used. This approach can be configured for existing workflows to detect other challenging and potentially underrepresented variants, thereby augmenting accurate variant calling in clinical NGS testing.

scholarly journals BAMscale: quantification of next-generation sequencing peaks and generation of scaled coverage tracks

2020 ◽  
Author(s):  
Lorinc Pongor ◽  
Jacob M Gross ◽  
Roberto Vera Alvarez ◽  
Junko Murai ◽  
Sang-Min Jang ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Replication Timing ◽  
Strand Break ◽  
Computational Time ◽  
Rna Seq ◽  
Next Generation ◽  
Dna Double Strand Break ◽  
Chromatin Immunoprecipitation Sequencing ◽  
One Step ◽  
Generation Sequencing

Abstract Background: Next-generation sequencing allows genome-wide analysis of changes in chromatin states and gene expression. Data analysis of these increasingly used methods either requires multiple analysis steps, or extensive computational time. We sought to develop a tool for rapid quantification of sequencing peaks, and an efficient method to produce coverage tracks for accurate visualization that can be intuitively interpreted by experimentalists with minimal bioinformatics background. We demonstrate its strength by integrating data from several types of sequencing approaches. Results: We have developed BAMscale, a one-step tool that processes a wide set of sequencing datasets. To demonstrate the usefulness of BAMscale, we analyzed multiple sequencing datasets from chromatin immunoprecipitation sequencing (ChIP-seq) data, chromatin state change data (Assay for Transposase-Accessible Chromatin using sequencing: ATAC-seq, DNA double strand break mapping sequencing: END-seq), DNA replication data (Okazaki fragments sequencing: OK-seq, Nascent-strand sequencing: NS-seq, single-cell replication timing sequencing: scRepli-seq) and RNA-seq data. The outputs consist of raw and normalized peak scores (multiple normalizations) in text format and scaled bigWig coverage tracks that are directly accessible to data visualization programs. Our tool can effectively analyze large sequencing datasets (~100Gb size) in minutes, outperforming currently available tools.Conclusions: BAMscale is a tool that can be used to accurately quantify and normalize identified peaks directly from BAM files, as well as create coverage tracks for visualization in genome browsers. BAMScale can be implemented for a wide set of methods for calculating coverage tracks such as ChIP-seq / ATAC-seq, as well as splice aware RNA-seq, END-seq and OK-seq for which no dedicated software is available. BAMscale is freely available on github (https://github.com/ncbi/BAMscale).

scholarly journals Highly sensitive and full-genome interrogation of SARS-CoV-2 using multiplexed PCR enrichment followed by next-generation sequencing

2020 ◽  
Author(s):  
Chenyu Li ◽  
David N. Debruyne ◽  
Julia Spencer ◽  
Vidushi Kapoor ◽  
Lily Y. Liu ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Multiplex Pcr ◽  
High Sensitivity ◽  
Detection Methods ◽  
Next Generation ◽  
Rt Pcr ◽  
Copy Numbers ◽  
Virus Rna ◽  
Highly Sensitive ◽  
Generation Sequencing

AbstractMany detection methods have been used or reported for the diagnosis and/or surveillance of COVID-19. Among them, reverse transcription polymerase chain reaction (RT-PCR) is the most commonly used because of its high sensitivity, typically claiming detection of about 5 copies of viruses. However, it has been reported that only 47-59% of the positive cases were identified by some RT-PCR methods, probably due to low viral load, timing of sampling, degradation of virus RNA in the sampling process, or possible mutations spanning the primer binding sites. Therefore, alternative and highly sensitive methods are imperative. With the goal of improving sensitivity and accommodating various application settings, we developed a multiplex-PCR-based method comprised of 343 pairs of specific primers, and demonstrated its efficiency to detect SARS-CoV-2 at low copy numbers. The assay produced clean characteristic target peaks of defined sizes, which allowed for direct identification of positives by electrophoresis. We further amplified the entire SARS-CoV-2 genome from 8 to half a million viral copies purified from 13 COVID-19 positive specimens, and detected mutations through next generation sequencing. Finally, we developed a multiplex-PCR-based metagenomic method in parallel, that required modest sequencing depth for uncovering SARS-CoV-2 mutational diversity and potentially novel or emerging isolates.

scholarly journals A highly sensitive method for the detection of recombinant PERV-A/C env RNA using next generation sequencing technologies

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Ken Kono ◽  
Kiyoko Kataoka ◽  
Yuzhe Yuan ◽  
Keisuke Yusa ◽  
Kazuhisa Uchida ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Endogenous Retrovirus ◽  
High Titre ◽  
Next Generation ◽  
Sequencing Technologies ◽  
Porcine Islet ◽  
Highly Sensitive ◽  
Generation Sequencing ◽  
Sensitive Method

AbstractSeveral xenogenic cell-based therapeutic products are currently under development around the world for the treatment of human diseases. Porcine islet cell products for treating human diabetes are a typical example. Since porcine cells possess endogenous retrovirus (PERV), which can replicate in human cells in vitro, the potential transmission of PERV has raised concerns in the development of these products. Four subgroups of infectious PERV have been identified, namely PERV-A, -B, -C, and recombinant PERV-A/C. Among them, PERV-A/C shows a high titre and there was a paper reported that an incidence of PERV-A/C viremia was increased in diseased pigs; thus, it would be important to monitor the emergence of PERV-A/C after transplantation of porcine products. In this study, we developed a highly sensitive method for the detection of PERV-A/C using next generation sequencing (NGS) technologies. A model PERV-C spiked with various doses of PERV-A/C were amplified by RT-PCR and the amplicons were analysed by NGS. We found that the NGS analysis allowed the detection of PERV-A/C at the abundance ratios of 1% and 0.1% with true positive rates of 100% and 57%, respectively, indicating that it would be useful for the rapid detection of PERV-A/C emergence after transplantation of porcine products.

Next-generation sequencing identifies a novelELAVL1–TYK2fusion gene in MOLM-16, an AML cell line highly sensitive to the PIM kinase inhibitor AZD1208

2016 ◽  
Vol 57 (12) ◽  
pp. 2927-2929 ◽  
Author(s):  
Adriana E. Tron ◽  
Erika K. Keeton ◽  
Minwei Ye ◽  
Matias Casas-Selves ◽  
Huawei Chen ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Cell Line ◽  
Kinase Inhibitor ◽  
Next Generation ◽  
Highly Sensitive ◽  
Generation Sequencing
Sign in / Sign up

Export Citation Format

Share Document