In the first quarter of 2016, we presented a NGS webinar series in which our experts discussed the applications of NGS technology, starting from sample to developing incredible insights. We got a lot of questions from the audience and in this article we share the top 9 frequently asked questions with in-depth responses from our team. Hope you enjoy it! Let us know if you have any additional questions or feel free to share your views in the comment section.
1. With which NGS platforms is your target gene amplification kit compatible?
The GeneRead DNAseq Targeted Panels V2 are compatible with the current Illumina® and Ion Torrent® sequencers on the market. Following target enrichment, you can construct the NGS library using the GeneRead DNA Library Core I Kit (for Illumina) or L Kit (for Ion Torrent), depending on the NGS platform.
2. Will this technology be used in the future for pre-symptomatic detection as a screening test?
QIAseq and GeneRead Library Prep Kits are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.
3. What is the minimum read depth you would suggest when dealing with liquid biopsy cfDNA with targeted DNA sequencing?
We recommend a read depth of 1500x for variant calling of somatic mutations.
4. How different is this approach than capture-based methods for enrichment? Which approach should I use?
Capture-based methods, such as on-array and in-solution hybrid capture, is suitable when your sequencing target is large. PCR-based target enrichment is used when the target is around 1 Mb. So, your approach will depend on your research needs. The PCR-based method is more specific (93% vs 80%); more uniform (80% vs 61%) and more reproducible (100% vs 95%). Here is a reference: http://bit.ly/1V9QBUF
5. What types of variants can you detect with this enrichment approach? Is this for human DNA only?
You can detect point mutations, indels, and copy number variants
6. Can I build a panel for hotspots only?
Yes, you can use the GeneRead DNAseq Custom Builder to build fully customized panels for targeted enrichment using multiplex PCR.
7. How much DNA can I get from a liquid biopsy?
One can get about 1–50 ng of circulating cell free DNA from 1 ml of plasma. The yield varies because a lot depends not only on the nature of the sample (cancer, non-cancer etc) but also on how the sample was collected, stored and processed.
- 8. Do amplicons usually cover all exon regions of a gene or just some hotspots?
Both. The majority of panels cover exons, while the human tumor actionable mutation panel covers hotspots.
9. Can you comment on comparison of your panels with Illumina‘s TruSight® Tumor 15 or 26 or Cancer panels?
TruSight Tumor 15 provides assessment of 15 genes commonly mutated in solid tumors. We offer the highest number and largest range of predesigned target enrichment panels, including 2 panels for solid tumors, 1 for hematology malignancy, 9 for specific cancers, 1 for specific genes (BRCA1 and BRCA2) and 2 comprehensive cancer panels. The number of genes in these panels varies from 8 in the Tumor Actionable Mutation panel to 160 in the Comprehensive Cancer panel, depending on the panel type.
In case you missed the webinar series, here are the links to the recordings of the “Meet the NGS expert’s” webinars:
Webinar 1: NGS: a review of its impact on test development, by Dr. Birgit Funke Link
Webinar 2: NGS to identify relevant mutations in cfDNA, by Dr. Clifford Tepper Link
Webinar 3: Clinical sequencing system to classify meningioma, by Dr. Nishihara Link
Trademarks: Illumina®, TruSight® (Illumina, Inc.); Ion Torrent® (Thermo Fisher)