Screening by single-molecule molecular inversion probes targeted sequencing panel of candidate genes of infertility in azoospermic infertile Jordanian males

2021 ◽  
pp. 1-8
Author(s):  
Osamah Batiha ◽  
George J. Burghel ◽  
Ayesha Alkofahi ◽  
Emad Alsharu ◽  
Hannah Smith ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Single Molecule ◽  
Targeted Sequencing ◽  
Molecular Inversion Probes

scholarly journals Comprehensive identification of pathogenic gene variants in patients with neuroendocrine disorders

2021 ◽  
Author(s):  
Sebastian Alexis Vishnopolska ◽  
Maria Florencia Mercogliano ◽  
Maria Andrea Camilletti ◽  
Amanda Helen Mortensen ◽  
Debora Braslavsky ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Single Molecule ◽  
Genetic Screening ◽  
Genetic Diagnosis ◽  
Incomplete Penetrance ◽  
Promoter Activation ◽  
Pathogenic Variants ◽  
Uncertain Significance ◽  
Molecular Inversion Probes ◽  
Congenital Hypopituitarism

Abstract Purpose Congenital hypopituitarism (CH) can present in isolation or with other birth defects. Mutations in multiple genes can cause CH, and the use of a genetic screening panel could establish the prevalence of mutations in known and candidate genes for this disorder. It could also increase the proportion of patients that receive a genetic diagnosis. Methods We conducted target panel genetic screening using single-molecule molecular inversion probes sequencing to assess the frequency of mutations in known hypopituitarism genes and new candidates in Argentina. We captured genomic DNA from 170 pediatric patients with CH, either alone or with other abnormalities. We performed promoter activation assays to test the functional effects of patient variants in LHX3 and LHX4. Results We found variants classified as pathogenic, likely pathogenic or with uncertain significance in 15.3% of cases. These variants were identified in known CH causative genes (LHX3, LHX4, GLI2, OTX2 and HESX1), in less frequently reported genes (FOXA2, BMP4, FGFR1, PROKR2, PNPLA6) and in new candidate genes (BMP2, HMGA2, HNF1A, NKX2-1). Conclusion In this work, we report the prevalence of mutations in known CH genes in Argentina and provide evidence for new candidate genes. We show that CH is a genetically heterogeneous disease with high phenotypic variation and incomplete penetrance, and our results support the need for further gene discovery for CH. Identifying population-specific pathogenic variants will improve the capacity of genetic data to predict eventual clinical outcomes.

scholarly journals Identification of community-consensus clinically relevant variants and development of single molecule molecular inversion probes using the CIViC database

2018 ◽  
Author(s):  
Erica K. Barnell ◽  
Adam Waalkes ◽  
Kelsi Penewit ◽  
Katie M. Campbell ◽  
Zachary L. Skidmore ◽  
...  
Keyword(s):  
Single Molecule ◽  
Pearson Correlation ◽  
Genomic Region ◽  
Variant Annotation ◽  
Design Paradigm ◽  
Molecular Inversion Probes ◽  
Sequencing Platforms ◽  
Tumor Types ◽  
Molecular Inversion Probe ◽  
Actionable Variants

AbstractClinical targeted sequencing panels are important for identifying actionable variants for cancer patients, however, there are currently no strategies to create impartial and rationally-designed panels to accommodate rapidly growing knowledge within the field. Here we use the Clinical Interpretations of Variants in Cancer database (CIViC) in conjunction with single-molecule molecular inversion probe (smMIP) capture to identify and design probes targeting clinically relevant variants in cancer. In total, 2,027 smMIPs were designed to target 111 eligible CIViC variants. The total genomic region covered by the CIViC smMIPs reagent was 61.5 kb. When compared to existing genome or exome sequencing results (n = 27 cancer samples from 5 tumor types), CIViC smMIP sequencing demonstrated a 95% sensitivity for variant detection (n = 61/64 variants). Variant allele frequency for variants identified on both sequencing platforms were highly concordant (Pearson correlation = 0.885; n = 61 variants). Moreover, for individuals with paired tumor/normal samples (n = 12), 182 clinically relevant variants missed by original sequencing were discovered by CIViC smMIPs sequencing. This design paradigm demonstrates the utility of an open-sourced database built on attendant community contributions for each variant with peer-reviewed interpretations. Use of a public repository for variant identification, probe development, and variant annotation could provide a transparent approach to build a dynamic next-generation sequencing–based oncology panel.

BRCA Testing by Single-Molecule Molecular Inversion Probes

2017 ◽  
Vol 63 (2) ◽  
pp. 503-512 ◽  
Author(s):  
Kornelia Neveling ◽  
Arjen R Mensenkamp ◽  
Ronny Derks ◽  
Michael Kwint ◽  
Hicham Ouchene ◽  
...  
Keyword(s):  
Genetic Testing ◽  
Copy Number Variation ◽  
Single Molecule ◽  
Copy Number ◽  
Single Gene ◽  
Work Flow ◽  
Analytical Sensitivity ◽  
Brca1 And Brca2 ◽  
Number Variation ◽  
Molecular Inversion Probes

Abstract BACKGROUND Despite advances in next generation DNA sequencing (NGS), NGS-based single gene tests for diagnostic purposes require improvements in terms of completeness, quality, speed, and cost. Single-molecule molecular inversion probes (smMIPs) are a technology with unrealized potential in the area of clinical genetic testing. In this proof-of-concept study, we selected 2 frequently requested gene tests, those for the breast cancer genes BRCA1 and BRCA2, and developed an automated work flow based on smMIPs. METHODS The BRCA1 and BRCA2 smMIPs were validated using 166 human genomic DNA samples with known variant status. A generic automated work flow was built to perform smMIP-based enrichment and sequencing for BRCA1, BRCA2, and the checkpoint kinase 2 (CHEK2) c.1100del variant. RESULTS Pathogenic and benign variants were analyzed in a subset of 152 previously BRCA-genotyped samples, yielding an analytical sensitivity and specificity of 100%. Following automation, blind analysis of 65 in-house samples and 267 Norwegian samples correctly identified all true-positive variants (>3000), with no false positives. Consequent to process optimization, turnaround times were reduced by 60% to currently 10–15 days. Copy number variants were detected with an analytical sensitivity of 100% and an analytical specificity of 88%. CONCLUSIONS smMIP-based genetic testing enables automated and reliable analysis of the coding sequences of BRCA1 and BRCA2. The use of single-molecule tags, double-tiled targeted enrichment, and capturing and sequencing in duplo, in combination with automated library preparation and data analysis, results in a robust process and reduces routine turnaround times. Furthermore, smMIP-based copy number variation analysis could make independent copy number variation tools like multiplex ligation-dependent probes amplification dispensable.

Abstract 4894: Targeted sequencing of myeloid cancers using single-molecule enrichment in picodroplets

2015 ◽  
Author(s):  
Michael L. Samuels ◽  
Steve K. Kotsopoulos ◽  
Frances Long ◽  
Holly Gettler ◽  
Omo Clement ◽  
...  
Keyword(s):  
Single Molecule ◽  
Targeted Sequencing

scholarly journals Targeted sequencing of candidate genes of dyslipidemia in Punjabi Sikhs: Population-specific rare variants in GCKR promote ectopic fat deposition

PLoS ONE ◽  
2019 ◽  
Vol 14 (8) ◽  
pp. e0211661 ◽  
Author(s):  
Dharambir K. Sanghera ◽  
Ruth Hopkins ◽  
Megan W. Malone-Perez ◽  
Cynthia Bejar ◽  
Chengcheng Tan ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Rare Variants ◽  
Fat Deposition ◽  
Targeted Sequencing ◽  
Ectopic Fat ◽  
Ectopic Fat Deposition

scholarly journals Conventional and Single-Molecule Targeted Sequencing Method for Specific Variant Detection in IKBKG while Bypassing the IKBKGP1 Pseudogene

2018 ◽  
Vol 20 (2) ◽  
pp. 195-202 ◽  
Author(s):  
Glynis Frans ◽  
Wim Meert ◽  
Jutte Van der Werff Ten Bosch ◽  
Isabelle Meyts ◽  
Xavier Bossuyt ◽  
...  
Keyword(s):  
Single Molecule ◽  
Targeted Sequencing ◽  
Sequencing Method ◽  
Variant Detection

scholarly journals Single molecule targeted sequencing for cancer gene mutation detection

2015 ◽  
Author(s):  
Yan Gao ◽  
Liwei Deng ◽  
Qin Yan ◽  
Yongqian Gao ◽  
Zengding Wu ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Gene Mutation ◽  
Single Molecule ◽  
Mutation Detection ◽  
Hla Typing ◽  
Targeted Sequencing ◽  
Rapid Decline ◽  
Cancer Gene ◽  
Cancer Genes ◽  
Targeted Capture

With the rapid decline cost of sequencing, it is now clinically affordable to examine multiple genes in a single disease-targeted test using next generation sequencing. Current targeted sequencing methods require a separate step of targeted capture enrichment during sample preparation before sequencing, and the library preparation process is labor intensive and time consuming. Here, we introduced an amplification-free Single Molecule Targeted Sequencing (SMTS) technology, which combined targeted capture and sequencing in one step. We demonstrated that this technology can detect low-frequency mutations of cancer genes. SMTS has several advantages, namely that it requires little sample preparation and avoids biases and errors introduced by PCR reaction. This technology can be applied in cancer gene mutation detection, inherited condition screening and high-resolution human leukocyte antigen (HLA) typing.

scholarly journals Full-Length Transcriptome Analysis Reveals Candidate Genes Involved in Terpenoid Biosynthesis in Artemisia argyi

2021 ◽  
Vol 12 ◽  
Author(s):  
Yupeng Cui ◽  
Xinqiang Gao ◽  
Jianshe Wang ◽  
Zengzhen Shang ◽  
Zhibin Zhang ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Single Molecule ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Enrichment Analysis ◽  
Full Length ◽  
Functional Enrichment ◽  
Terpenoid Biosynthesis ◽  
Sequencing Data ◽  
Artemisia Argyi

Artemisia argyi is an important medicinal plant widely utilized for moxibustion heat therapy in China. The terpenoid biosynthesis process in A. argyi is speculated to play a key role in conferring its medicinal value. However, the molecular mechanism underlying terpenoid biosynthesis remains unclear, in part because the reference genome of A. argyi is unavailable. Moreover, the full-length transcriptome of A. argyi has not yet been sequenced. Therefore, in this study, de novo transcriptome sequencing of A. argyi's root, stem, and leaf tissues was performed to obtain those candidate genes related to terpenoid biosynthesis, by combining the PacBio single-molecule real-time (SMRT) and Illumina sequencing NGS platforms. And more than 55.4 Gb of sequencing data and 108,846 full-length reads (non-chimeric) were generated by the Illumina and PacBio platform, respectively. Then, 53,043 consensus isoforms were clustered and used to represent 36,820 non-redundant transcripts, of which 34,839 (94.62%) were annotated in public databases. In the comparison sets of leaves vs roots, and leaves vs stems, 13,850 (7,566 up-regulated, 6,284 down-regulated) and 9,502 (5,284 up-regulated, 4,218 down-regulated) differentially expressed transcripts (DETs) were obtained, respectively. Specifically, the expression profile and KEGG functional enrichment analysis of these DETs indicated that they were significantly enriched in the biosynthesis of amino acids, carotenoids, diterpenoids and flavonoids, as well as the metabolism processes of glycine, serine and threonine. Moreover, multiple genes encoding significant enzymes or transcription factors related to diterpenoid biosynthesis were highly expressed in the A. argyi leaves. Additionally, several transcription factor families, such as RLK-Pelle_LRR-L-1 and RLK-Pelle_DLSV, were also identified. In conclusion, this study offers a valuable resource for transcriptome information, and provides a functional genomic foundation for further research on molecular mechanisms underlying the medicinal use of A. argyi leaves.

scholarly journals Single Molecule Molecular Inversion Probes for High Throughput Germline Screenings in Dystonia

2019 ◽  
Vol 10 ◽  
Author(s):  
Michaela Pogoda ◽  
Franz-Joachim Hilke ◽  
Ebba Lohmann ◽  
Marc Sturm ◽  
Florian Lenz ◽  
...  
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Molecular Inversion Probes
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