Liu, H., Jian, L., Xu, J., Zhang, Q., Zhang, M., Jin, M., Peng, Y., Yan, J., Han, B., Liu, J., Gao, F., Liu, X., Huang, L., Wei, W., Ding, Y., Yang, X., Li, Z., Zhang, M., Sun, J., Bai, M., Song, W., Chen, H., Sun, X., Li, W., Lu, Y., Liu, Y., Zhao, J., Qian, Y., Jackson, D., Fernie, A.R., Yan, J. (May 2020) High-Throughput CRISPR/Cas9 Mutagenesis Streamlines Trait Gene Identification in Maize. Plant Cell, 32 (5). pp. 1397-1413. ISSN 1040-4651
Abstract
Maize is one of the most important crops in the world. However, few agronomically important maize genes have been cloned and used for trait improvement, due to the complex genome and genetic architecture. Here we integrated multiplexed CRISPR/Cas9-based high-throughput targeted mutagenesis with genetic mapping and genomic approaches to successfully knock-out 743 candidate genes corresponding to agronomic and nutritional traits. After low-cost barcode-based deep sequencing, 412 edited sequences covering 118 genes were precisely identified from individuals showing clear phenotypic changes. This mutant profile was similar to the ones identified in human cell lines and predictable. An unexpectedly frequent homology-directed repair through endogenous templates was observed and likely caused by the spatial contact between distinct chromosomes. Through several case studies on validation and interpretation of gene function, this targeted mutagenesis library suggested that the integration of forward- and reverse- genetics promises rapid validation of important agronomic genes for crops with complex genomes. Beyond specific findings, this study also guides further optimization of high-throughput CRISPR experiments in plants.
Actions (login required)
 |
Administrator's edit/view item |