Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

Next-generation Sequencing03:00

Next-generation Sequencing

92.3K
The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features....
92.3K
Sanger Sequencing01:57

Sanger Sequencing

756.1K
DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...
756.1K
Modern Molecular Taxonomy01:29

Modern Molecular Taxonomy

91
Advancements in molecular biology have revolutionized the identification and characterization of bacteria, with multiple methods leveraging DNA sequencing for enhanced precision. As sequencing technologies improve and costs decline, these approaches are increasingly used in clinical, environmental, and evolutionary studies.Multilocus Sequence Typing (MLST) examines several housekeeping genes, essential chromosomal genes encoding cellular functions, to distinguish strains. Approximately...
91
RNA-seq03:21

RNA-seq

10.3K
RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases. 
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while...
10.3K

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Analysis of purines and pyrimidines across biospecimens: influence of storage temperature and duration.

Scientific reports·2026
Same author

Sensitivity of HiFi long-read genome sequencing for difficult-to-detect pathogenic variants when applied to real-world clinical laboratory samples.

American journal of human genetics·2026
Same author

Implementation of First-Line Rapid Genome Sequencing for Children in Pediatric and Cardiac Intensive Care Units.

American journal of medical genetics. Part A·2026
Same author

Multiscore, a gene ranker powered by artificial intelligence and real-world clinical data, shows high sensitivity for the molecular diagnosis of Mendelian disorders in nearly 10,000 exomes and genomes.

Human genetics·2026
Same author

Clinical Insights Into Disaccharidase Deficiency: Prevalence, Intestinal Histology, and Outcomes.

Mayo Clinic proceedings·2025
Same author

From Expert Knowledge to Validation Resources: A Case for Using in Silico Approaches to Close the Gap in Available Reference Materials for Common Germline Genetic Tests.

The Journal of molecular diagnostics : JMD·2025

Related Experiment Video

Updated: Aug 25, 2025

Detection of Rare Genomic Variants from Pooled Sequencing Using SPLINTER
14:06

Detection of Rare Genomic Variants from Pooled Sequencing Using SPLINTER

Published on: June 23, 2012

15.3K

Scalable detection of technically challenging variants through modified next-generation sequencing.

Susan Rojahn1, Tina Hambuch1, Jessika Adrian1

  • 1Invitae, San Francisco, California, USA.

Molecular Genetics & Genomic Medicine
|October 17, 2022
PubMed
Summary

Next-generation sequencing (NGS) methods were adapted to accurately detect challenging genetic variants, including those with high-similarity copies or repetitive sequences. These NGS adaptations offer a scalable and cost-effective approach for comprehensive genetic variant identification.

Keywords:
bioinformaticsgenetic testingmolecular geneticsnext-generation sequencing

More Related Videos

Rare Event Detection Using Error-corrected DNA and RNA Sequencing
10:36

Rare Event Detection Using Error-corrected DNA and RNA Sequencing

Published on: August 3, 2018

12.2K
Detection of Rare Mutations in CtDNA Using Next Generation Sequencing
11:11

Detection of Rare Mutations in CtDNA Using Next Generation Sequencing

Published on: August 24, 2017

16.9K

Related Experiment Videos

Last Updated: Aug 25, 2025

Detection of Rare Genomic Variants from Pooled Sequencing Using SPLINTER
14:06

Detection of Rare Genomic Variants from Pooled Sequencing Using SPLINTER

Published on: June 23, 2012

15.3K
Rare Event Detection Using Error-corrected DNA and RNA Sequencing
10:36

Rare Event Detection Using Error-corrected DNA and RNA Sequencing

Published on: August 3, 2018

12.2K
Detection of Rare Mutations in CtDNA Using Next Generation Sequencing
11:11

Detection of Rare Mutations in CtDNA Using Next Generation Sequencing

Published on: August 24, 2017

16.9K

Area of Science:

  • Genetics
  • Bioinformatics
  • Molecular Biology

Background:

  • Certain clinically significant genetic variants pose challenges for standard next-generation sequencing (NGS) due to high-similarity copies, repetitive sequences, and other complex genomic structures.
  • Examples of such challenging variants include those in PMS2, SMN1/SMN2, GBA1, HBA1/HBA2, CYP21A2, ARX, FMR1, CFTR, and MSH2.

Purpose of the Study:

  • To develop and validate customized NGS processes capable of detecting technically challenging genetic variants.
  • To improve the sensitivity and specificity of NGS for a range of complex genetic markers.

Main Methods:

  • Customization of NGS protocols, incorporating target enrichment strategies.
  • Implementation of bioinformatic masking techniques to address highly similar sequences.
  • Validation of adapted NGS methods using samples with known genotypes.

Main Results:

  • The adapted NGS assays demonstrated high sensitivity (100%) for all tested technically challenging variants.
  • High specificity was achieved for most variants (e.g., PMS2, GBA1, SMN1/SMN2, HBA1/HBA2, MSH2 Boland inversion), with slightly lower but still high specificity for CYP21A2 and ARX variants.
  • The approach provides a sensitive primary assay, with options for orthogonal disambiguation when necessary.

Conclusions:

  • Refined NGS chemistries and bioinformatics enable the detection of previously challenging genetic variants.
  • The described adaptations offer a scalable and cost-effective method for identifying all clinically relevant genetic variants within a single sample.