| Actin is a ubiquitous component of the plant cytoskeleton and participates in a number of important subcellular processes.The microfilament cytoskeleton is involved in cell division plane localization,cell elongation and cell shape determination.The plant actins are made up of 376 or 377 amino acids and they are highly conservative.The high conservativeness accords with the actins as function of cytoskeleton.High-quality RNA was extracted from Malus robusta seedlings.Then the cDNA was obtained through reverse transcription kit.An actin gene was obtained by rapid amplification of cDNA ends(RACE) from Malus robusta seedlings.Sequences analysis showed that the actin gene is 1134 base pair(bp),which encodes 377 amino acids.Sequences analysis showed there are only two different amino acids between MrACT and Arabidopsis thaliana actin 7.The predicted isoelectric point of the protein is 5.16.We detected that the expression of MrACT had no obvious difference in five kinds of tissues through RT-PCR assays.The result demonstrated that Malus robusta actin be used as internal standard to study expression of other internal genes in Malus robusta.Hydrogen peroxide(H2O2) was one of the reactive oxygen species(ROS) produced rapidly in the early stage of plant-pathogen interaction.When plant was infected by microbial pathogen,H2O2 played an important role in many defense responses that followed. So the plant's capability of broad-spectrum pathogen-resistance could be enhanced if H2O2 level was improved by gene transformation methods.Glucose oxidase could catalyze the oxidation ofβ-D-glucose to produce gluconic acid and H2O2.This enzyme had been detected in several bacteria and fungi,but hadn't been found in animals and plants.In order to study the GO gene more clearly,we constructed plant binary vectors.Then the plant binary vectors containing the GO gene which was controlled by CAMV35S promoter were constructed,they were transformed into Arabidopsis using Agrobacteriun-mediated transformation.Twelve transgenic Arabidopsis plants were obtained after agrobacterium-mediated transformation.GUS-staining and PCR analysis confirmed that the GO gene had integrated into the Arabidopsis genome,semi-quantitative RT-PCR revealed that the GO gene was transcripted in Arabidopsis plants.We detected the content of H2O2 in transgenic plants was higher than in wild type control plants.We also discovered that the content of flavonoids in transgenic plants was higher than in wild type control plants.Based on the advantage that the plant defense responses induced by H2O2 had no species restriction,it could be anticipated that the transgenic Arabidopsis plants acquired would have satisfied resistance to many pathogens.The accomplished test showed that the transgenic Arabidopsis plants had strong resistance to Pseudomonas(DC3000).Then the plant binary vectors containing the GO gene were constructed,they were transformed into Malus robusta.M using Agrobacteriun-mediated transformation. |
