AUTHOR=Park Jae-Ryoung , Kim Eun-Gyeong , Jang Yoon-Hee , Jan Rahmatullah , Farooq Muhammad , Ubaidillah Mohammad , Kim Kyung-Min 

TITLE=Applications of CRISPR/Cas9 as New Strategies for Short Breeding to Drought Gene in Rice

JOURNAL=Frontiers in Plant Science

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2022.850441

DOI=10.3389/fpls.2022.850441

ISSN=1664-462X

ABSTRACT=<p>Recent unpredictable climate change is the main reason for the decline in rice yield. In particular, drought stress is a major constraint in reducing yield and quality for rice at rainfed agriculture areas, such as Asia and South America. CRISPR/Cas9 provides an effective solution for gene function study and molecular breeding due to specific editing of targeted genome sequences. In addition, CRISPR/Cas9 application can significantly reduce the time required to develop new cultivars with improved traits compared to conventional complex and time-consuming breeding. Here, drought-induced gene <italic>Oryza sativa Senescence-associated protein</italic> (<italic>OsSAP</italic>) was edited by CRISPR/Cas9. To investigate the possible role of <italic>OsSAP</italic> in drought stress, genome-editing plants were subjected to drought stress until the soil moisture content reached 20%, and the reactive oxygen species (ROS) scavenging efficiency of genome-editing plants were decreased. When the genome-editing plants were subjected to drought stress, survival rate, shoot length, root length, content of chlorophyll number of tiller, and 1,000-grain weight decreased, and more H<sub>2</sub>O<sub>2</sub> and O<sub>2</sub><sup>−</sup> were detected in leaves. In addition, expression levels of several critical stress-related transcription factors were decreased in the <italic>OsSAP</italic> genome-editing plant. These results suggest that <italic>OsSAP</italic> function as a positive regulator during drought stress response in rice. We analyzed the expression of <italic>OsSAP</italic> and <italic>Cas9</italic> in T<sub>0</sub> and T<sub>1</sub> plants as well as T<sub>2</sub> seeds. As the course of generation advancement progressed, <italic>Cas9</italic> expression remained stable or weakened but the <italic>OsSAP</italic> expression was continuously removed from the T<sub>0</sub> plant. The coefficient of variation (CV) in both T<sub>1</sub> plants and T<sub>2</sub> seeds was lower than 5%. Overall, our results suggest that CRISPR/Cas9 could be a novel and important tool for efficiently generating specific and inheritable targeted genome editing in rice, with short breeding cycles.</p>