Research Of The Physiological Changes Of Bougainvillea Under Salt Stress And Cloning Of SOD Genes From Bougainvillea

Bougainvillea (Bougainvillea spp.) is an important landscape tree species in southern China, in this study, an annual cuttage seedling of purple Bougainvillea and pink Bougainvillea were respectively treated with different salt concentration. The rate of electrolyte leakage, MDA, proline, protective enzymes and other physiological changes were determined in leaves of the 2 species of Bougainvillea in salt stress, to explore the change and difference of Bougainvillea resistant index in salt stress; liquid chromatography method were used for the determination of some endogenous hormones in the Bougainvillea leaves, to study the relationship between the changes of hormone content in the process of salt stress; at the same time, the Bougainvillea SOD gene family were cloned by the method of RACE, provides the molecular basis for the study of resistance of Bougainvillea spectabilis. The main results are as follows:1. Annual of purple Bougainvillea and pink Bougainvillea cuttage seedling as materials,2 varieties of Bougainvillea were respectively treated in 0.2%-0.6% salt solution. The results show that the membrane stability, protective enzyme activities and contents of osmotic adjustment substances in Bougainvillea leaf changes with the treatment time and concentration. The electrolyte leakage, MDA, proline content of pink Bougainvillea leaves were increased significantly, and increased with the increase of time and concentration of salt, the soluble protein content increased first and then decreased, SOD activity of POD increased, CAT activity decreased; purple Bougainvillea is shown as the electrolyte leakage, MDA, proline content increasing, and increases with the rising time and salt concentration, the content of soluble protein increased first and then decreased, the SOD activity of POD first increased and then decreased, CAT activity decreased. In the process of stress, the purple Bougainvillea is sensitive and the rise and fall of physiological changes is bigger change than pink Bougainvillea. Therefore, electrolyte leakage, MDA, proline and the triangle plum is closely related to salt tolerance.2. Liquid chromatographic method were used to determin the changes hormone content of Bougainvillea leaf in the process of salt stress. The content of ABA in leaves of pink Bougainvillea increased under salt stress, salt treatment for 0.4% and 0.6% were significant increased, GA3 content was first increased then decreased, IAA content decreased; The changes of ABA, GA3 and IAA content in is similar to the pink Bougainvillea. GA3 in 0.6% salt treatment of purple Bougainvillea is greater than the pink, and morphological features as the majority of leaves fallen. So that the change of GA3 content is closely related to salt tolerance of Bougainvillea.3. Reference homology cloning method, cDNA of Bougainvillea different types of SOD gene family were cloned by using RACE technology, the results are as follows:Clone of CSD (Cu/ZnSOD):Cytoplasmic CSD complete sequences were obtained by a total of 8, including 4 types of cyCSD1, with the same ORF,585bp in length, encoding a protein of 195 amino acids,3’UTR length ranging from 147bp-237bp, length of 5’UTR is 55bp; 4 types of cyCSD2, with the same ORF,543bp in length, encoding 180 amino acid,3’UTR length ranging from 147bp-237bp, length of 5’UTR was 43bp. The chloroplastic CSD, only part of the conserved region of 428bp long fragment was cloned. Through the Blast online comparison analysis, cyCSD and spinach cytoplasmic Cu/ZnSOD is 84% similarity, Mesembryanthemum crystallinum cytoplasmic Cu/ZnSOD as 82% similarity, chCSD and Chenopodium murale chloroplast Cu/ZnSOD similarity 89%, and Cottonwood chloroplast Cu/ZnSOD as 87% similarity, that is both the Cu/ ZnSOD sequence of bougainvillea.Clone of FSD(FeSOD):The chloroplastic FSD fragments were obtained as 841bp. By the analysis of online blast alignment, and similarity with grape and longan FSD gene respectively reached 85% and 78% landed in Genbank, considered that is the FSD sequence of bougainvillea.Clone of MSD (MnSOD):Three complete mitochondrial MSD sequences were cloned, with the same ORF, as 711bp length, encoding a protein of 236 amino acids,3 ’UTR was 282bp-314bp length,5’UTR length was 72bp. By the analysis of online blast comparison, with similarity of Salicornia and Suaeda salsa MSD landed in Genbank were respectively as 84% and 83%, considered as the MSD sequence of bougainvillea.