Screening and functional verification of drought resistance-related genes in castor bean seeds

Abstract Due to its limited freshwater resources, China has large arid and semiarid regions. Drought is one of the natural stresses that greatly impact plants. Castor bean (Ricinus communis L.) is an oil crop with high economic value. Drought is one of the factors limiting castor bean growth. The drought resistance mechanisms of castor bean have become a research focus. In this study, we screened out the genes related to drought resistance in castor bean embryos at the germination stage through combined physiological measurements、proteomic and metabolomic analysis. These genes were heterologously overexpressed and complementarily expressed in Arabidopsis thaliana to analyze their roles in the response mechanism of A. thaliana to drought resistance. The experimental results are as follows: The germination stage embryos of castor oil treated with water for 48 h (S48), 60 h (S60), and 15% PEG 6000 for 48 h (P48) and 60 h (P60) were used as experimental materials, and the embryos of S48 and S60 were used as experimental materials. As a control, the physiological activity (CAT, SOD, POD, GST, T-AOC), content (MDA, H2O2, Pro) and hydroxyl radical scavenging ability of embryo samples P48 and P60 treated with 15% PEG 6000 for the same time were measured. The results showed that compared with the control, with the increase of treatment time, the CAT activity, SOD activity, POD activity, H2O2 content, Pro content and hydroxyl radical scavenging ability of castor embryos all showed an upward trend, while GST activity, T-AOC activity and T-AOC activity increased. The activity and MDA content showed a downward trend. Based on combined physiological measurements、proteomic and metabolomic analysis and RT-qPCR validation, three significantly upregulated genes, RcECP63 (LOC8274875), RcDDX31 (LOC8282119), and RcA/HD1 (LOC8277163), were finally selected for study. We compared the growth, germination, expression of RcECP63, RcDDX31, and RcA/HD1, and physiological indicators of four types (mutant, complementary-expression, wild-type, and overexpression) of A. thaliana under drought stress. The comparison showed that as the mannitol concentration increased, the A. thaliana plants on mannitol medium exhibited shorter root lengths, less lateral roots, and lower germination rates. Compared with the controls, the drought-stressed overexpression and complementary-expression transgenic plants showed an overall trend of upregulation of RcECP63, RcDDX31, and RcA/HD1. The functions of the three genes in drought stress response were further verified by the measurement of physiological indicators.