Overexpression of Nicotiana tabacum High-affinity Potassium Ion Transporter Protein Gene(HAK1) Improves the Salt-tolerance in Tobacco(N. tabacum) Plants
1, 1, 1, 2
1. 2. Guizhou Key Lab of Agricultural Bioengineering,Guizhou University
Abstract：The high-affinity potassium ion transporter protein (HAK) plays an important role in improving the salt-tolerance in plants. In this study, we investigated the effect of salt stress on germination rate (Gr), germination potential (Gp), and germination index (Gi) in seeds of NtHAK1-overexpression plants and on the content of chlorophyll (Chl) and malondialdehyde (MDA), the activity of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) in NtHAK1-overexpressing tobacco plantlets (4 to 5 leaf stage). The results showed there was no siginificant effect on Gr, Gp, and Gi in the seeds of overexpressing and wild-type tobaccos under 50 mmol/L NaCl, but siginificant difference appeared between overexpressing plants and wild-type plants with 81.6% for Gr, 85.0% for Gp, and 42.6% for Gi in transgenic tobaccos under 100 mmol/L NaCl and 63.9% for Gr, 78.8% for Gp, and 20.2% for Gi in transgenic tobaccos under 150 mmol/L NaCl, which both were significantly higher than that in wild-type tobaccos (P＜0.05). The Chl content decreased from 2.25 mg/g (FW) before salt treatment to 1.31 mg/g(FW) at 5 d after salt treatment in wild-type plantlets and the decreasing amplitude of Chl content was 41.8%, while the average value of decreasing amplitude was 36.5% in transgenic tobaccos, which was significantly lower than the former. At 5 d after salt treatment, the MDA content was 105 nmol/g(FW) in non-transgenic plants, while it was 96.52 nmol/g(FW) in overexpressing plants. Meanwhile, the activities of antioxidant enzymes (AOEs), including SOD and CAT, were significantly up-regulated in NtHAK1-overexpression tobaccos with the increasing amplitude of 38.8% for SOD activity and 58.1% for CAT activity, while it was 34.2% for SOD and 54.6% for CAT in wild-type ones. The results from quantitative Real-time PCR for Na+/K+ absorption-related genes at 3 d after salt treatment in overexpressing and wild-type tobacco showed the relative expression levels of HAK1, potassium outward rectifier channel protein (TORK1), and vacuolar Na+/H+ antiporter protein (NHX1) were significantly up-regulated in NtHAK1- overexpression tobacco plants compared with wild-type ones. They were 3.85 folds for HAK1, 1.79 folds for TORK1, and 1.69 folds for NHX1 higher than those in wild-type tobaccos. In addition, the ratio of Na to K in different plant tissues demonstrated that it was 0.110 in root tissue and 0.106 in leaf tissue of transgenic plants respectively, while it was 0.147 for root tissue and 0.135 for leaf tissue in wild-type plants. The ratio of Na to K of different tissues in transgenic tobaccos was significantly lower (P＜0.05) than that in wild-type ones. This study provids the plant materials for breeding the salt-tolerant tobaccos and further studying the mechanism of HAK1-mediated salt-tolerance in transgenic plants.
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