|作者：||Gao Qingsong, Li Gang, Sun Hui, Xu Ming, Wang Huanhuan, Ji Jianhui, Wang Di, Yuan Caiyong, Zhao Xiangxiang|
1Jiangsu Collaborative Innovation Center of Regional Modern Agriculture & Environmental Protection/Jiangsu Key Laboratory for Eco-Agricultural Biotechnology around Hongze Lake, Huaiyin Normal University, Huai'an 223300, China.
2Huaiyin Institute of Agricultural Sciences of Xuhuai Region in Jiangsu, Huai'an 223001, China.
|刊名：||International journal of molecular sciences, 2020, Vol.21 (3)|
|关键词：||CRISPR/Cas9; FW2.2-like gene; Genome editing; Grain yield; Off-target effect; Rice; Tiller number;|
|原始语种摘要：||The FW2.2 -like ( FWL ) genes encode cysteine-rich proteins with a placenta-specific 8 domain. They play roles in cell division and organ size control, response to rhizobium infection, and metal ion homeostasis in plants. Here, we target eight rice FWL genes using the CRISPR/Cas9 system delivered by Agrobacterium -mediated transformation. We successfully generate transgenic T0 lines for 15 of the 16 targets. The targeted mutations are detected in the T0 lines of all 15 targets and the average mutation rate is found to be 81.6%. Transfer DNA (T-DNA) truncation is a major reason for the failure of mutagenesis in T0 plants. T-DNA segregation analysis reveals that the T-DNA inserts in transgenic plants can be easily eliminated in the T1 generation.... Of the 30 putative off-target sites examined, unintended mutations are detected in 13 sites. Phenotypic analysis reveals that tiller number and plant yield of OsFWL4 gene mutants are significantly greater than those of the wild type. Flag leaves of OsFWL4 gene mutants are wider than those of the wild type. The increase in leaf width of the mutants is caused by an increase in cell number. Additionally, grain length of OsFWL1 gene mutants is higher than that of the wild type. Our results suggest that transgene-free rice plants with targeted mutations can be produced in the T1 generation using the Agrobacterium -mediated CRISPR/Cas9 system and that the OsFWL4 gene is a negative regulator of tiller number and plant yield.|