Screening and Verification of Aquaporin Gene <i>AsPIP1-3</i> in Garlic (<i>Allium sativum</i> L.) under Salt and Drought Stress

Abstract

In order to screen candidate aquaporin genes involved in resisting osmotic stress, we analyzed the physiological responses and the expression levels of aquaporin genes in garlic under drought and salt stress with ‘Er Shuizao’ as plant material. Different physiological indicators were detected under drought and salt stress treatments. RT-qPCR was used to detect the expression levels of the candidate aquaporin genes in specific tissues. Finally, we screened AsPIP1-3 as a candidate gene and analyzed its function. The results showed that the relative water content and chlorophyll content of leaves decreased, the O2− production rate increased, and H2O2 accumulated in garlic under drought and salt stress. The activities of SOD, POD, and CAT enzymes first increased and then decreased in garlic. The content of soluble sugar and proline increased to maintain cell osmotic balance, and the content of MDA and relative conductivity continued to increase. Most aquaporin gene expression first increased and then decreased in garlic under drought and salt stress. AsPIP1-3 gene expression is up-regulated under drought and salt stress in garlic. The relative expression was the highest on the 6th day of stress, being related to antioxidant enzyme activity and osmotic regulation. The consistent changes in gene expressions and physiological responses indicated that AsPIP1-3 played a role in resisting garlic osmotic stress. AsPIP1-3 was located on the cell membrane, being consistent with the predicted results of subcellular localization. The germination rate and root length of transgenic Arabidopsis under drought stress were significantly different from the wild type. Drought stress reduced the ROS accumulation of transgenic Arabidopsis, and the antioxidant enzyme activity was significantly higher than the wild type. The relative conductivity and MDA content significantly decreased, and the proline content increased under drought stress. The expression level of the genes related to drought stress response (AtRD22, AtP5CS, AtABF3, and AtLEA) significantly increased. The overexpression of AsPIP1-3 genes improved the drought tolerance of transgenic Arabidopsis plants, showing that AsPIP1-3 proteins enhanced drought tolerance. Our study laid a foundation for exploring the regulatory mechanism of garlic to abiotic stress.

Keywords

• aquaporin
• osmotic stress
• expression analysis
• transgene
• functional verification