Yes, the TruSeq Amplicon - Cancer Panel has been optimized for use with DNA derived from FFPE sources. A simple qPCR-based quality control test (WG-321-1001, Illumina FFPE QC Kit) is recommended to ensure maximum sample success.
Yes, the TruSeq Amplicon - Cancer Panel has been optimized for use with DNA derived from FFPE sources. The input amount depends on the ∆Cq. For more information, see the TruSeq FFPE DNA Library Prep QC Reference Guide (part # 15067391).
You can use the Illumina FFPE QC Assay (WG-321-1001) to determine whether your samples meet the minimum quality criteria for inclusion in a Cancer Panel experiment. This protocol is qPCR based, where a Ct (threshold cycle) cut-off allows you to determine whether your samples will succeed in the TruSeq Amplicon assay or not. This measurement represents how well DNA amplifies and can be used to infer DNA quality. Testing has demonstrated that over 80% of samples passing the Illumina FFPE QC Assay protocol are expected to succeed in the TruSeq Amplicon assay.
Use the Qiagen QIAamp DNA FFPE Tissue Kit (catalog # 56404).
The cancer panel is an off-the-shelf standard, or fixed, panel that cannot be customized. TruSeq Custom Amplicon offers the ability to fully customize targets of interest using DesignStudio.
The smallest supported batch size is 16 samples (15 samples plus one control). The 16-sample minimum accounts for up to six freeze/thaw cycles of the 96-sample kit.
For MiSeq Reagent Kits v2 300-cycle configuration, we recommend that up to 60 samples per MiSeq run are pooled. When this guidance is followed, >1000x mean coverage per amplicon per sample should be achieved in runs with 15M clusters passing filter. The MiSeq Reagent Kits v3 can deliver >25M clusters passing filter and can accommodate 96 cancer panel samples per MiSeq run to achieve the same >1000x mean depth of coverage.
The TruSeq Amplicon - Cancer panel is fully supported on the MiSeq System. This panel takes advantage of the long read lengths (2 x 150 bp) and the MiSeq integrated software solution. TruSeq Custom Amplicon and the TruSeq Amplicon – Cancer Panel are compatible with the HiSeq 2500 System.
The manifest file is available for download from MyIllumina.
The TruSeq Amplicon - Cancer Panel is a presynthesized, fixed content panel. As such, the content cannot be modified. However, the genomic targets from the cancer panel can be used as a starting point for a customized TruSeq Custom Amplicon design using DesignStudio.
Mutational hotspots from over 48 genes associated with cancer are targeted with 212 amplicons.
The TruSeq Amplicon - Cancer Panel requires a 2 x 150 bp read length. This read length allows bidirectional sequencing of amplicons in the panel.
No. File directory structures from a HiSeq system are incompatible with MiSeq Reporter software.
However, the TruSeq Amplicon App is available in BaseSpace and can be used to analyze the this kit.