Targeted Genotyping of a Whole-Gene Repertoire by an Ultrahigh-Multiplex and Flexible HD-Marker Approach
Pingping Liu,
Jia Lv,
Cen Ma,
Tianqi Zhang,
Xiaowen Huang,
Zhihui Yang,
Lingling Zhang,
Jingjie Hu,
Shi Wang,
Zhenmin Bao
Affiliations
Pingping Liu
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China; Laboratory for Marine Biology and Biotechnology, Pilot Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, China
Jia Lv
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China; Corresponding authors.
Cen Ma
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China
Tianqi Zhang
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China
Xiaowen Huang
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China
Zhihui Yang
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China
Lingling Zhang
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China; Laboratory for Marine Biology and Biotechnology, Pilot Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, China
Jingjie Hu
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China; Laboratory for Marine Fisheries Science and Food Production Processes, Pilot Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, China
Shi Wang
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China; Laboratory for Marine Biology and Biotechnology, Pilot Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, China; Laboratory for Marine Fisheries Science and Food Production Processes, Pilot Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, China
Zhenmin Bao
Key Laboratory of Marine Genetics and Breeding and Sars-Fang Center, (Ministry of the People’s Republic of China) Ocean University of China, Qingdao 266003, China; Laboratory of Tropical Marine Germplasm Resources and Breeding Engineering, Sanya Oceanographic Institution, Ocean University of China, Sanya 572000, China; Laboratory for Marine Fisheries Science and Food Production Processes, Pilot Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, China; Corresponding authors.
Targeted genotyping is an extremely powerful approach for the detection of known genetic variations that are biologically or clinically important. However, for non-model organisms, large-scale target genotyping in a cost-effective manner remains a major challenge. To address this issue, we present an ultrahigh-multiplex, in-solution probe array-based high-throughput diverse marker genotyping (HD-Marker) approach that is capable of targeted genotyping of up to 86 000 loci, with coverage of the whole gene repertoire, in what is a 27-fold and six-fold multiplex increase in comparison with the conventional Illumina GoldenGate and original HD-Marker assays, respectively. We perform extensive analyses of various ultrahigh-multiplex levels of HD-Marker (30 k-plex, 56 k-plex, and 86 k-plex) and show the power and excellent performance of the proposed method with an extremely high capture rate (about 96%) and genotyping accuracy (about 96%). With great advantages in terms of cost (as low as 0.0006 USD per genotype) and high technical flexibility, HD-Marker is a highly efficient and powerful tool with broad application potential for genetic, ecological, and evolutionary studies of non-model organisms.
