MicroRNAs (miRNAs) are small non-coding RNAs with roles in plant development
and stress responses. Loss-of-function analysis of miRNA genes has been
challenging due to the lack of suitable knockout tools. A team of scientists
from various universities, led by Jian-Ping Zhou from the University of
Electronic Science and Technology of China, aim to study miRNA genes,
specifically OsMIR528, in rice using CRISPR-Cas9..
Frequencies of mutants T0 lines ranged from 48% to 89% at all target sites.
Three independent guide RNAs (gRNAs) all generated biallelic mutations among
mutant lines. This demonstrates that CRISPR-Cas9 is an effective tool for
knocking out plant miRNAs. However, single-base pair (bp) mutations in
mature miRNA regions were found to lead to the generation of functionally
redundant miRNAs, while large deletions were found to abolish miRNA
function. Analysis found that OsMIR528 is a positive regulator of salt
This work provides guidelines on targeting miRNAs with CRISPR-Cas9 and also
brings new insights into miRNA function in rice.