Cloning of the Soybean <i>GmNHL1</i> Gene and Functional Analysis under Salt Stress

When encountered in the soybean seedling stage, salt stress has serious impacts on plant growth and development. This study explores the role of the soybean <i>NDR1/HIN1-like</i> family gene <i>GmNHL1</i> under salt stress. First, the <i>GmNHL1</i> gene was successfully cloned, and bioinformatic analysis revealed multiple cis-acting elements which are related to adversity stress and involved in the oxidative response in the promoter region. Sub-cellular localization analysis indicated that the protein expressed by <i>GmNHL1</i> was localized on the cell membrane. An over-expression vector of the target gene and a CRISPR/Cas9 gene-editing vector were constructed, and the recipient soybean variety Jinong 74 was genetically transformed using the <i>Agrobacterium tumefaciens</i>-mediated method. By analyzing the performance of the different plants under salt stress, the results showed that <i>GmNHL1</i> was over-expressed in the T2 generation. The germination potential, germination rate, germination index, and vitality index of the strain were significantly higher than those of the recipient control JN74. Under salt stress conditions, the root microanatomical structure of the <i>GmNHL1</i> over-expressing material remained relatively intact, and its growth was better than that of the recipient control JN74. Measurement of physiological and biochemical indicators demonstrated that, compared with the receptor control JN74, the malondialdehyde and O₂⁻ contents of the <i>GmNHL1</i> over-expressing material were significantly reduced, while the antioxidant enzyme activity, proline content, and chlorophyll content significantly increased; however, the results for <i>GmNHL1</i> gene-edited materials were the opposite. In summary, over-expression of <i>GmNHL1</i> can improve the salt tolerance of plants and maintain the integrity of the root anatomical structure, thereby more effectively and rapidly reducing the accumulation of malondialdehyde and O₂⁻ content and increasing antioxidant enzyme activity. This reduces cell membrane damage, thereby improving the salt tolerance of soybean plants. These results help to better understand the mechanism of salt tolerance in soybean plants, laying a theoretical foundation for breeding new stress-resistant varieties of soybean.