Effects Of AtTIP And AtNHX On Salt Tolerance Of Arabidopsis Thaliana

Cultivation of salt-tolerant crops through genetic engineering is an important way to developing the saline. Predecessors and previous studies in our laboratory found that the corresponding gene expression of the Na + / H + exchanger and the tonoplast intrinsic protein rise in euhalophytes under saline conditions. Therefore, we specμlated that these genes are probably involved in salt tolerance of plants. In this article, one tonoplast intrinsic protein(tonoplast intrinsic proteins, TIP1-1) and two Na+/H+exchangers(Na+/H+ exchanger, NHX) gene were overexpressed respectively in Arabidopsis. Then the role of these three genes in salt tolerance of Arabidopsis was compared. The main results that we obtained are as follows.1 At NHX1、At NHX2、At TIP1-1gene clone and transform Arabidopsis thaliana The CDS sequence of At NHX1, At NHX2 and At TIP1-1 were respectively cloned through RT-PCR method. Use p CAMBIA-35s-OCS and p Cambia1300-221-HA-MYC as vector backbone, the CDS sequence of the three genes were cloned into it, and construct At NHX1, At NHX2 and At TIP1-1 gene overexpression vector. Then the overexpression vector was transformed Arabidopsis through floral dipping method mediated by Agrobacterium. By the means of normal PCR and quantitative PCR to detect Kanamycin-resistant and hygromycin-resistant plants, and found that all of transgenic plants can transcript and express of the target gene normaly.2 Transgenic Arabidopsis morphological analysis under different salt concentrations When the transgenic and wild-type Arabidopsis germinated to two cotyledons,then transfered them to square plate of MS medium containing0,50,100,150 and 200 m M Na Cl. 7 days later, it turned out that the salt-resistance of the At NHX1 and At NHX2 Arabidopsis lines were obviously better than the wild type at 100-200 m M Na Cl concentration.However,the salt-resistance of At TIP1-1Arabidopsis lines were not as good as wild-type Arabidopsis at the concentration of150-200 m M Na Cl, which were called salt-sensitive.Transgenic plants and the wild-type Arabidopsis were not significantly different at the other salt concentrations.3 Transgenic Arabidopsis physiological and biochemical analysis under salt treatment When transgenic Arabidopsis grown to six-leaf stage, plants were treated with100 m M and 200 m M Na Cl for two weeks. Then chlorophyll fluorescence,chlorophyll content, fresh weight and protein content were measured. The results showed that Fv/Fm, chlorophyll content and fresh weight of salt-treated 35s: At NHXs Arabidopsis were all higher than that of WT Plants, which indicated that 35s: At NHX2 Arabidopsis had similar salt-tolerance with 35s: At NHX1, which are important to salttolerance in plants. But, At TIP1-1 can not improve salt-tolerance in Arabidopsis, and even more sensitive to salt stress. All results illustrated that the At TIP1-1 may collaborate with At NHX to regulate plants salt tolerance in the tonoplast. This synergy between the Na+ /H+ exchanger and the tonoplast intrinsic protein will be proved in the future.