Abstract
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 Oryza sativa Senescence-associated protein (OsSAP) was edited by CRISPR/Cas9. To investigate the possible role of OsSAP 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 H2O2 and O2− were detected in leaves. In addition, expression levels of several critical stress-related transcription factors were decreased in the OsSAP genome-editing plant. These results suggest that OsSAP function as a positive regulator during drought stress response in rice. We analyzed the expression of OsSAP and Cas9 in T0 and T1 plants as well as T2 seeds. As the course of generation advancement progressed, Cas9 expression remained stable or weakened but the OsSAP expression was continuously removed from the T0 plant. The coefficient of variation (CV) in both T1 plants and T2 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.
Highlights
To respond to rapid climate change, it is necessary to improve cultivars of crops and develop rice with resistance to biotic and abiotic stress; this is a primary goal in plant breeding worldwide (Zhang et al, 2018)
In the CRISPR/Cas9 vector, which can be expressed in rice, the Oryza sativa Senescence-associated protein (OsSAP) guide RNA is regulated by the U3 promoter (Figure 1A)
By applying CRISPR/Cas9, the functional loss of OsSAP interfered with the antioxidant defense by reducing the activation of superoxide dismutase (SOD), POD, and CAT, and the negative effects of various agricultural traits due to the accumulation of reactive oxygen species (ROS) and MDA resulted in leaf aging and reduction 1,000-grain weight
Summary
To respond to rapid climate change, it is necessary to improve cultivars of crops and develop rice with resistance to biotic and abiotic stress; this is a primary goal in plant breeding worldwide (Zhang et al, 2018). Recent rapid increases in the global temperature have raised awareness of the fragility of the world’s food supply, and solutions are needed (Siebert et al, 2017). It has recently been established that drought stress is the main cause of the reduction of rice yield, and drought can have a direct impact on future food shortages (Passioura, 2007; Prabnakorn et al, 2018). One specific problem in relation to drought is how to maintain stable yield, while minimizing the drought-related effects of climate change on various food crops. Plants undergo many structural and physiological changes during drought; the breeding of drought stress-resistant crops has become a means by which to strengthen the plants (Yang et al, 2019)
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