Confidence is the currency of science.
IDT will be participating in European Society for Human Genetics (ESHG) 2020 virtually this year. Learn why top sequencing companies choose IDT and why IDT scores high in customer service—especially for our NGS products.
- Enabling variant calling in challenging FFPE samples by coupling a novel library preparation chemistry with exome sequencing
This study demonstrates that the xGenTM Exome Research Panel v2, when combined with xGen Prism DNA library preparation, provides researchers with a complete human exome FFPE-sequencing solution with robust performance across FFPE samples of varying quality.
Though targeted next generation sequencing (NGS) is widely applied in genetic diagnostics and research, achieving high sensitivity presents challenges, such as detecting low levels of mutant sequences or target RNA molecules due to a low fraction of aberrant cells.
Our first presentation reviews two studies: 1) the detection of gene-doping using a DNA-based NGS panel and 2) the measurement of minimal residual disease (MRD), the persistence of leukemic cells after treatment, and using an RNA-based NGS panel. Our second presentation highlights a sequencing protocol for SARS-CoV-2 based on the established amplicon sequencing approach known as the ‘ARTIC method’.
- Detection of low levels of mutant sequences with NGS
Presented by: Eddy de Boer
University of Groningen, University Medical Center Groningen, Department of Genetics
Targeted NGS effectiveness is challenged when high sensitivity is required—for example, in the detection of low levels of mutant sequences or when the presence of the target RNA molecule is low due to a low fraction of aberrant cells. Here are two studies aimed at overcoming such challenges: 1) detection of gene doping  using a DNA-based NGS panel and 2) measuring minimal residual disease (MRD) , the persistence of leukemic cells after treatment, using an RNA-based NGS panel.
1) Existing PCR-based methods for gene doping detection in athletes target exon-exon junctions in the intron-less transgene, but evasion is possible by tampering with cDNA sequences. We designed tamper-resistant xGen Lockdown Probes directed at all exon-exon junctions of 5 potential doping genes. All junctions were detectable with a sensitivity of 1296 cDNA copies in 1000 ng of genomic DNA and plasmid-derived sequences were observable; optimizations are possible.
2) Measuring MRD is important for monitoring leukemia recurrence; current monitoring uses flow cytometry and digital droplet PCR (ddPCR). NGS advantages include the ability to analyze different genetic aberrations and patients in 1 experiment. We designed a) xGen Lockdown Probes targeting the breakpoints of 13 fusion transcripts; b) non-labeled probes blocking wild-type transcripts; and c) xGen Lockdown Probes fully tiling housekeeping genes for quantification. Our NGS method reached a maximum sensitivity of 1 aberrant cell per 10,000 and was mostly within a factor 10 compared to ddPCR. Further optimizations are easy and will likely boost sensitivity.
Explore some of our NGS solutions
xGen Prism DNA Library Prep Kit
xGen Exome Research Panel v2
NGS Discovery Pools
xGen Lockdown Probe Pools
xGen Lockdown Panels
xGen Blocking Oligos
Custom NGS Adapters