Knocking Out MicroRNA Genes in Rice with CRISPR-Cas9
Background: MicroRNAs (miRNAs) are small noncoding RNAs that play important roles in plant development and stress responses. Loss-of-function analysis of miRNA genes has been traditionally challenging due to lack of appropriate knockout tools. In this chapter, we describe a method of using CRISPR-Cas9 for knocking out microRNA genes in rice by Agrobacterium-mediated transformation. We also demonstrate single-strand conformation polymorphism (SSCP) as an effective genotyping method for screening CRISPR-Cas9-induced mutations.
Key wordsMicroRNAs CRISPR-Cas9 Genome editing Rice SSCP
This work was supported by grants including the National Science Foundation of China (31330017 and 31771486), the Sichuan Youth Science and Technology Foundation (2017JQ0005), the National Transgenic Major Project (2018ZX08022001-003), the Fundamental Research Funds for the Central Universities (ZYGX2016J119 and ZYGX2016J122) to YZ and XZ, and a start-up fund from the University of Maryland, College Park to YQ.
- 18.Zhou J, Deng K, Cheng Y, Zhong Z, Tian L, Tang X, Tang A, Zheng X, Zhang T, Qi Y, Zhang Y (2017) CRISPR-Cas9 based genome editing reveals new insights into MicroRNA function and regulation in rice. Front Plant Sci 8:1598. https://doi.org/10.3389/fpls.2017.01598CrossRefPubMedPubMedCentralGoogle Scholar