Cloning And Expression Of GST Gene Of Suaedu Salsa And Transformation Of Suaeda CDNA Over-expression Library In Arabidopsis

alt stress consists of osmotic and ionic stress, then the secondary stress which is called oxidative stress following the formers. The imposition of environmental stress leads to increasing production of reactive oxygen species (ROS) in plant cells, which can damage proteins, membrane lipids. DNA and other cellular components. Plants have evolved protection mechanisms which include enzymatic reaction system and no-enzymatic reaction system. The enzymatic reaction system mainly includes superoxide dismutase (SOD), catalase(CAT), gluiathione S-transferase(GST). glutathione peroxidase(GOPX), ascorbate peroxidase(APX) and so on. Glutathione S-transferases can not only play a fundamental role in protection agaist endogenous or exogenous toxic chemicals but also directly scavenge ROS of the cells efficiently with glutathione peroxidase activity.The first cDNA clone for gluiathione S-iransferase (GST) in plants was isolated from corns, in the following from soybean, potato, tabacco and Arahidopsis ihulianu. It has been confirmed that overexpression of GST in iransgenic tabacco effectively has increased tolerance of plant to salt stress and cold stress.Suaeda salsa, a kind of plant characteristic of seawatcr-lulerant. can grow normally under 400 mmol/L NaCl. An EST about 0.6kb has been obtained among more than 1000 Expressed Sequence Tags (ESTs) from young seedlings of .V. salsa by constructing a cDNA library. According to the results of homeologous analysis, the ESTwas shown to be 55% homologous with the GST cDNA of Glycine max reported earlier. The full length sequence of the EST was obtained by bidirectional sequencing, the GenBank number is AF321437. Sequencing analysis showed that the cDNA probably encodes glutahtione S-transferase enzyme consisted of 235 amino acids. Compared the sequence of amino acids with the four GST cDNA which are higher homologous, it shows the relatively higher conservation. Results of Southern blotting suggest that there may be more than two copies in genomic DNA of .S1. salsa. Northern blotting shows that the relative expression levels under 400rmnol/L NaCl for 48 h are as 2-3 times high as the control, which indicates GST gene in S.salsa is salt-induced.We have developed a system to over-express cDNA library of Suueda salsa, and it was used to transform to Arabic/apsis thalianu on a large scale. We constructed an overexpression vector consisted of the part of pCAMB!A1200 and pRT!05. The Suuccki cDNA library were driven by the cauliflower mosaic virus (CaMV) 35S promoter. The cDNA library were identified using PCR with the flanking primers, showing that at least 94% of this library have cDNA inserts. The pooled plasmids were introduced into Arabidopsis upon Agrobacterium tumefciens-mediated transformation by Floral-dip methods. About 500 individual Hygromycin-B resistant plants were obtained from 100.000 seeds. Eight of transgenic plants have some different phenotypes from wild type plants. Only 20% of TI plants have one inserts. PCR checking indicated that the expression vector had been integrated into the transgenic plants" genomes. The analysis of salt tolerance of transgenic plants is now in process.