PT - JOURNAL ARTICLE AU - Veronica Gonzalez AU - Sivaraman Natarajan AU - Yuntao Xia AU - David Klein AU - Robert Carter AU - Yakun Pang AU - Bridget Shaner AU - Kavya Annu AU - Daniel Putnam AU - Wenan Chen AU - Jon Connelly AU - Shondra Pruett-Miller AU - Xiang Chen AU - John Easton AU - Charles Gawad TI - Accurate Genomic Variant Detection in Single Cells with Primary Template-Directed Amplification AID - 10.1101/2020.11.20.391961 DP - 2020 Jan 01 TA - bioRxiv PG - 2020.11.20.391961 4099 - http://biorxiv.org/content/early/2020/11/20/2020.11.20.391961.short 4100 - http://biorxiv.org/content/early/2020/11/20/2020.11.20.391961.full AB - Improvements in whole genome amplification (WGA) would enable new types of basic and applied biomedical research, including studies of intratissue genetic diversity that require more accurate single-cell genotyping. Here we present primary template-directed amplification (PTA), a new isothermal WGA method that reproducibly captures >95% of the genomes of single cells in a more uniform and accurate manner than existing approaches, resulting in significantly improved variant calling sensitivity and precision. To illustrate the new types of studies that are enabled by PTA, we developed direct measurement of environmental mutagenicity (DMEM), a new tool for mapping genome-wide interactions of mutagens with single living human cells at base pair resolution. In addition, we utilized PTA for genome-wide off-target indel and structural variant detection in cells that had undergone CRISPR-mediated genome editing, establishing the feasibility for performing single-cell evaluations of biopsies from edited tissues. The improved precision and accuracy of variant detection with PTA overcomes the current limitations of accurate whole genome amplification, which is the major obstacle to studying genetic diversity and evolution at cellular resolution.Competing Interest StatementCG is a co-founder and board member of BioSkryb, which is commercializing primary template-directed amplification.