3.4.3. Overexpression of CaHSP18.1a Enhances Plant Tolerance to Salt Stress

To study the role of *CaHSP18.1a* in salt stress, transgenic *Arabidopsis thaliana* was subjected to salt stress. First, the germination rate of transgenic seeds under salt stress was observed (Supplementary Figure S3A). OE2, OE1, and WT seeds exhibited normal germination on MS plates without NaCl; the germination rate and seedling growth of transgenic lines were almost unaffected by 100 mM NaCl MS plates. However, the seed germination rate differed when the NaCl concentration was increased to 150 mM, and the germination rate of WT seeds was slightly lower than that of transgenic seeds. The salt tolerance of *CaHSP18.1a* transgenic *Arabidopsis* seeds increased. After the germinated seedlings were moved to MS plates with NaCl concentrations of 0, 100, and 150 mM, compared with the untreated seedlings, the growth of OE2, OE3, OE1, and WT seedlings after 6 d under 100 and 150 mM NaCl treatments was worse (Supplementary Figure S3A,B); root elongation was significantly inhibited, and the root length of transgenic lines seedlings was greater than that of WT plants (Figure 8A).

**Figure 6.** Drought resistance of transgenic *CaHSP18.1a*-OE *Arabidopsis* plants. (**A**) Phenotypes of wild-type (WT) and *CaHSP18.1a*-OE *Arabidopsis*; (**B**,**C**) Malonaldehyde (MDA) content and relative electrolyte leakage (REL) of WT and *CaHSP18.1a*-OE *Arabidopsis*; (**D**–**G**) Superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and ascorbic acid peroxidase (APX) activity of WT and *CaHSP18.1a*-OE *Arabidopsis* without watering for 10 days. Data are means with standard deviations of three biological replicates. Different letters denote statistical significance (*p* ≤ 0.05).

**Figure 7.** Expression pattern of stress–response genes in wild-type (WT), OE1, OE2, and OE3 lines before and after 10 days of drought treatment. Data are means with standard deviations of three biological replicates. Different letters denote statistical significance (*p* ≤ 0.05).

**Figure 8.** Overexpression of *CaHSP18.1a* enhanced tolerance to salt stress. (**A**) Root length of wild-type (WT) and *CaHSP18.1a*-OE *Arabidopsis* lines grown for 10 d on Murashige and Skoog (MS) medium containing 0, 100, and 150 mM NaCl. (**B**) Seedling growth for WT and transgenic plants exposed to NaCl. Three-week-old seedlings were watered with 200 mM NaCl once every 1–2 days. The images were taken after 7 days.

To explore the effect of *CaHSP18.1a* on the salt tolerance of *Arabidopsis thaliana*, 3 week-old WT and transgenic plants were irrigated with 200 mM NaCl solution for 7 days. As shown in Figure 8B, WT plants showed dehydration and wilting with weak growth. However, the transgenic *Arabidopsis thaliana* showed no significant difference from the control plants except for a slight yellowing phenotype in their leaves.
