Thellungiella Glutathione Peroxidase Gene Cloning And Expression Analysis

Glutathione peroxidase (GPX) is one of the crucial enzymes which play an important part in cleaning reactive oxidative species (ROS) in plant. From now on, systematic research on GPXs can be done in Arabidopsis thaliana, Lotus japonicus belonging to leguminous plants, and Populus trichocarpa belonging to woody plantThGPXs gene in Thellungiella salsuginea and their sequences(in DNA level and protein level), expression patterns under salt and drought stresses were analyzed respectively. Moreover, the subcellular localization of ThGPX6was studied. The results we obtained were as follows:1. Based on the ESTs and transcriptome data, ThGPXs were cloned with electronic cloning and RACE technology, and named as ThGPXl~ThGPX8. According to the sequence analysis, ThGPXs had167-236amino acid residues and their molecular weight were18.9-26.3kDa. ThGPX2and ThGPX8were acid proteins, ThGPX3was neutral protein, while others were alkaline proteins.2. In genome research on ThGPXs, they shared the same gene structure in exon and intron arrangements and different UTR regions, and each UTR region occupies a single exon. Every ThGPXs member had6exons and5introns. The intron splicing site obeyed the GT-AG rule. The lengths of ThGPXs'exon were highly conserved. Three conserved motifs in GPX were located in the conserved exons. There were many different c/s-elements in their promoter regions, including light, hormone and abiotic stress responded elements.3. Bioinformatic research showed that, ThGPXs had many irregular curves as the main protein secondary structure and only had a little (3-sheet. C-terminuses were mainly consisted of a-helix. ThGPXs always form dimmers in vivo. We can also find obvious differences at a-helix in C-terminus. Among ThGPXs members, ThGPX6has the highest number of potential phosphorylation sites. Signal peptide and trans-membrane motif can only be found in ThGPX3, which was a secreting protein. Based on PSORT tool, ThGPX2distributed in cytoplasm. ThGPX3was located on cytoplasma membrane and endoplasmic membrane; ThGPX6was in mitochondria, while ThGPXl, ThGPX7, and ThGPX8were all chloroplast proteins.4. Real-time quantitative PCR results showed that, expression patterns of ThGPXs varied in different tissues under salt and osmotic stresses. Under treatment of300mM NaCl, ThGPXs were upregulated and reached the highest level at24h. Except ThGPX7, others in salt cress expressed in higher levels after48h treatment of20%PEG6000.5. Subcellular localization results displayed that ThGPX6was a mitochondria protein, which showed its function in cleaning ROS.Our studies have revealed the structure and function of ThGPXs to some extend. However, the mechanisms of GPXs in abiotic stress are still unclear. Additionally, plants can be stimulated under multiple stresses at the same time, for example, the drought stress followed by heat and light stress. Further study is still needed because of unresolved biological functions in ThGPXs under stress.