TELL-Seq-based phasing of Cas9-excised human BRCA1/2 fragments. Homo sapiens

Many clinical and research applications require phasing long haplotypes. To add long-range phasing capabilities to Illumina platforms, we have developed the Transposase Enzyme-Linked Long-read-Sequencing (TELL-Seq) method. TELL-Seq is a linked-read methodology that requires as little as 0.1 ng of input DNA, processed in a single tube in less than three hours. We have tested TELL-Seq for phasing of approximately 190 kb segments of DNA containing the BRCA1 or BRCA2 genes--both genes associated with an increased risk for breast and ovarian cancers. These two DNA fragments were excised and isolated from human cells (PCR-free) using CRISPR/Cas9 technology and the HLS-CATCH system. After TELL-Seq processing, phasing was conducted using a reference genome (GRCh38) or by de novo assembly-i.e., reference-free. Our results demonstrate high recall accuracy for variant annotations and phasing using Genome in a Bottle (GIAB) Personal Genome Project cells (B lymphocytes) for GM24385 (HG002), GM24149 (HG003), and GM24143 (HG004).