TruSight Cardio Sequencing Kit Training FAQs

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  • Analysis


  • TruSight Cardio regions, targets, and probes are provided in the *.bed file format.  The *.bed file can be imported into either the UCSC Genome Browser (genome.ucsc.edu) or the Integrated Genome Viewer (IGV) (www.broadinstitute.org/igv/).

    To create subpanels, generate gene lists in HGNC nomenclature for the regions of interest. Upload this subpanel gene list when importing VCF files into VariantStudio, and then filter variants based on the shortened gene list.

    For reference, see the complete TruSight Cardio gene list.

    For sequencing of TruSight Cardio libraries on the MiSeq, use the MiSeq Reporter Enrichment workflow or the Generate FASTQ workflow. Illumina recommends preparing the sample sheet in the Illumina Experiment Manager (IEM). Libraries prepared with TruSight Cardio Sequencing kit (MiSeq, 12 samples) are dual-indexed, but only require a single i7 Index Read for demultiplexing. Set up the run for paired-end 151-cycle reads and a single 8 bp Index Read. FASTQ files can be uploaded to BaseSpace and analyzed using the BWA Enrichment App or the Isaac Enrichment App (v2.0 and v2.1 custom manifest workflow). Alternatively, use the MiSeq Reporter Enrichment workflow. 

    When using BaseSpace, set up the run in BaseSpace Prep tab. Select Nextera Rapid Capture from the dropdown menu under Library Prep kit to associate samples with the indexes provided in the TruSight Cardio Sequencing kit (NextSeq, 48 samples, Mid-output), E502, E503, E505, E506, and N701-12. Set up the run as a dual index paired-end 151-cycle sequencing run. FASTQ files streamed into BaseSpace can be analyzed using the BWA Enrichment App or the Issac Enrichment App (v2.0 and v2.1 custom manifest workflow).

    When running the NextSeq in Standalone mode, enter the following parameters on the Run Setup Screen:

    Read Type: Paired End

    Read Length: Read 1: 151; Read 2: 151; Index 1: 8; Index 2: 8

    When the run is complete, use the bcl2fastq2 converter for demultiplexing and FASTQ conversion. Create a sample sheet for demultiplexing using Illumina Experiment Manager. Choose NextSeq; Nextseq FASTQ only; Nextera Rapid Capture Enrichment Sample Prep kit. Perform data analysis using third-party software.