OsmiR535 belongs to the miR156/miR529/miR535 superfamily, a highly conserved miRNA family in plants. OsmiR535 is involved in regulating the cold-stress response, modulating plant development, and determining panicle architecture and grain length. However, the role that OsmiR535 plays in plant responses to drought and salinity are elusive. In the current study, molecular and genetic engineering techniques were used to elucidate the possible role of OsmiR535 in response to NaCl, PEG(Poly ethylene glycol), ABA(Abscisic acid), and dehydration stresses. Our results showed that OsmiR535 is induced under stressed conditions as compared to control. With transgenic and CRISPR/Cas9 knockout system techniques, our results verified that either inhibition or knockout of OsmiR535 in rice could enhance the tolerance of plants to NaCl, ABA, dehydration and PEG stresses. In addition, the overexpression of OsmiR535 significantly reduced the survival rate of rice seedlings during PEG and dehydration post-stress recovery. Our results demonstrated that OsmiR535 negatively regulates the stress response in rice. Moreover, our practical application of CRISPR/Cas9 mediated genome editing created a homozygous 5 bp deletion in the coding sequence of OsmiR535, demonstrating that OsmiR535 could be a useful genetic editing target for drought and salinity tolerance and a new marker for molecular breeding of Oryza sativa.

MicroRNAs (miRNAs) are endogenous small non-coding RNAs that play an important role in post-transcriptional gene regulation in plants and animals by target

Transposable elements (TEs), also known as transposons, a type of mobile genetic elements, are widespread across all investigated eukaryotic organisms and typically constitute the major portion of most genomes, especially in grasses, where they can account for up to 90% of the genome (Vitte et al., 2014). They not only are actively involved in altering gene structure and regulating gene expression, but also have played a profound role in reshaping genomic architecture and maintaining genomic stability (Lisch, 2013).