Cloing Of GR2 Gene And Expression Analysis Of Antioxidant Enzyme Gene Under Adversity Stress In Brassica Campestris L.

The growth of plants are influenced by some non-biology factors such as under cold, high temperature, dry, salt stress and so forth.These effects have been caused enormous losses in agriculture by reducing harvest of crops.Recent years, there are increasing people making research about some genes of antioxidant enzyme.Brassica campestris L. is one of the most efficient oil crops during the world. The new rapeseed varieties Longyou6 has a high strength cold resistance, it is currently the only one can in Gansu Province, north-central He Xi regional security wintering of winter oilseed rape varieties. We did a preliminary study about the sequence features and functions of GR2 genes in Longyou 6 and Tianyou 2. RT-PCR method was used to analyze the expression of GR2 and five genes of antioxidant enzyme?GR1, SOD, CAT, APX, GPX?in different stress. This research results provide the theory evidence for more deeply research of anti-oxidant system.1. A new GR2 gene was isolated from Longyou 6. The full-length c DNA was 2073 bp contained a 1692 bp opening reading frame and comprising 563 amino acid residues with the predicted molecular weight 60.7 k D and isoelectric point 7.9. Senior predict protein structure show that GR2 rich in alpha helix, a hydrophilic protein. The protein had a total of 1-3 transmembrane regions and without signal peptide sequences. Comprehension of amino homology, phylogenetic tree and prediction location software, the result indicates GR2 gene has high homology with GR of many plants.They have low-level homology by comparing GR1 with GR2, and GR2 belongs to chloroplast GR in Brassica campestris.2. The Real-time quantitative PCR?RT-PCR? result showed that GR2 was expressed in the roots, stem, leaves in the two kinds of Brassica campestris, and there are differences in different tissues, the expression in leaf is highest.3. The antioxidant enzyme genes expression under abiotic stress were analyzed by RT-PCR. Cold, high temperature, dry, salt stress induced the expression of two kinds of Brassica campestris. In different stress, GR1 was highest expressed in Longyou 6 and GR2 is highest expressed in Tianyou 2. ABA pretreatment before the above four kinds of adversity stress that GR1 and GR2 increasing clearly, and this trend more obvious in Longyou 6. H₂O₂ pretreatment increased the transcription level of GR1 but reduced GR2. The expression of CU/Zn-SOD, CAT, cAPX and GPX are induced by above-mentioned stress but the expression levels is difference; ABA pretreatment can increase expression of antioxidant enzymes genes after under stress or normal condition, however, the rise trend of every enzyme is differ both of Longyou 6 and Tianyou2; H₂O₂ treatment independently can induce the expression of antioxidant enzyme genes, and H₂O₂ pretreatment before four kinds of non-biology stress that enhanced expression of CU/Zn-SOD, cAPX and GPX, but induced expression of CAT. The MAPKK inhibitor U0126 pretreatment can inhibit expression of antioxidant enzymes in Brassica campestris both normal growth and under stress. Above results indicate that the low temperature, high temperature, drought and salt stress can inducedthe expression of antioxidant enzymes gene, and the signal molecules of ABA, H₂O₂ as well as MAPK cascade pathway involved in regulation of stress tolerance process.