FIND-IT: Accelerated trait development for a green evolution
Søren Knudsen, Toni Wendt, Christoph Dockter, Hanne Cecilie Thomsen, Magnus Wohlfahrt Rasmussen, Morten Egevang Jørgensen, Qiongxian Lu, Cynthia Voss, Emiko Murozuka, Jeppe Thulin Østerberg, Jesper Harholt, Ilka Braumann, J.A. Cuesta-Seijo, Sandip M. Kale, Sabrina Bodevin, Lise Tang Petersen, Massimiliano Carciofi, Pai Pedas, Jeppe Opstrup Husum, Martin Toft Simmelsgaard Nielsen, Kasper Nielsen, Mikkel K. Jensen, Lillian A. Møller, Zoran Gojković, Alexander Striebeck, Klaus B. Lengeler, Ross T. Fennessy, Michael Katz, Rosa García Sánchez, Natalia Solodovnikova, Jochen Förster, Ole Olsen, Birger Lindberg Møller, Geoffrey B. Fincher, Birgitte Skadhauge
Abstract
Improved agricultural and industrial production organisms are required to meet the future global food demands and minimize the effects of climate change. A new resource for crop and microbe improvement, designated FIND-IT (Fast Identification of Nucleotide variants by droplet DigITal PCR), provides ultrafast identification and isolation of predetermined, targeted genetic variants in a screening cycle of less than 10 days. Using large-scale sample pooling in combination with droplet digital PCR (ddPCR) greatly increases the size of low–mutation density and screenable variant libraries and the probability of identifying the variant of interest. The method is validated by screening variant libraries totaling 500,000 barley ( Hordeum vulgare ) individuals and isolating more than 125 targeted barley gene knockout lines and miRNA or promoter variants enabling functional gene analysis. FIND-IT variants are directly applicable to elite breeding pipelines and minimize time-consuming technical steps to accelerate the evolution of germplasm.