Study On The Mechanism Of Induced Drought Tolerance And Disease Resistance By Bacillus Cereus AR156

Because they can not move themself, plants always encounter abiotic and biotic attack. Induced systemic tolerance (IST) against to abiotic stress can be primed by many abiotic and biotic materies. Between them plant growth-promoting rhizobacteria (PGPR) are the most petential, they colonize the roots of plants for a long time, maintain humidity of soil, improve soil structure and mineral nutrition absorbing, promote plants growth and enhance disease resistance. Most important they are secure to humen being and animals. In this study, we studied the mechanism of induced systemic tolerance against drought stress by PGPR AR156 and the PGPR combination, hoping it will be helpful for practical application.Besides, PGPR widely exist in the plant rhizosphere soil. They compete with the pathogens for nutrition, colonization situs, or by producing some antibiotic substances to stop the infection, and promoting the growth of plants. Besides, PGPR can enhance disease systemic resistence of plants through systemic acquired resistance (SAR) and induced systemic resistence (ISR). Many transcription factors function as regulators in salicylic acid (SA) dependent pathway and ethylene/jasmonic acid (ET/JA) dependent pathway. In this study we screened transcription factors that regarded as regulators in these two signal pathways and constructed recombinant vectors for future functional studies.1. Bacillus cereus AR156 Induced Systemic Drought Tolerance of TomatoIn order to study the mechanism of drought torelance induced by AR156, we tested physiological indexes, the antioxidant enzymes activities and the expression of cytosolic ascorbate peroxidase gene (cAPX), monodehydroascorbate reductase gene (MDHAR) and ribulose-1,5-bisphosphate carboxy/oxygenase (Rubisco) small subunit gene (rbcS). Results showed that after treated with culture supernate of AR156, relative water content of leaves was 82%, increased by 44%, recovery intension of roots was 0.34 mg g-1 h-1, increased by 100%, chlorophyll a, chlorophyll b and total chlorophyll were increased by 16%,20% and 20.2%, respectively, and the expression of rbcS maintained a stable high level, besides monodehydroascorbate (MDA) content was 2.2×10-3?mol g-1 Fw, decreased by 21.4%, relative electrical conductivity was 56.0%, decreased by 28.2%, compared with control treatment. What's more, the activities of superoxide dismutase (SOD), peroxidase (POD) and hydrogen peroxidase (CAT) were increased markedly after applying with the culture supernate of AR156, and the expression of cAPX and MDHAR were up-regulated at anaphase of drought stress. Moreover the survival rate of tomato 24 h after re-watering was increased markedly, compared with control group. These results suggested that the culture supernate of PGPR Bacillus cereus AR156 induced systemic drought tolerance of tomato.2. Bacillus cereus AR156 Combination Induced Systemic Drought Tolerance of CucumberOur previous work showed that PGPR AR156 combination BBS(Bacillus cereus AR156, Bacillus subtilis SM21, and Serratia sp. XY21, BBS for short) was a potential biopesticide. In the present study, we present data that cucumbers watered with BBS received a systemic tolerance to drought stress. Under drought stress, MDA content and relative electrical conductivity were decreased by 40% and 15% respectively, proline content and the recovery intension of roots were increased by 3.45-fold and 50% respectively, and chlorophyll a, chlorophyll b and total chlorophyll content were significantly increased by 25.9%,31.5% and 27.4% after BBS treatment, compared with control group. Besides, activity of superoxide dismutase (SOD) was much higher than that of control group, and the expression of cytosolic ascorbate peroxidase gene (cAPX), ribulose-1,5-bisphosphate carboxy/oxygenase (Rubisco) large subunit gene (rbcL) and small subunit gene (rbcS) decreased slower in cucumber leaves with BBS treated. However, none of the three strains of BBS produced 1-aminocyclopropane-l-carboxylate (ACC) deaminase. These results suggested that PGPR combination BBS was an elicitor of induced systemic tolerance (1ST) to drought stress in cucumber, although members of it couldn't degrade ethylene precursor ACC.3. Screening and Plant Expression Vector Construction of the Transcription Factors Regulating the Priming in Arabidopsis Induced by Bacillus cereus AR156Previous studied showed that Bacillus cereus AR156 induced systematic resistance by priming SA-dependent pathway and ET/JA -dependent pathway. In order to investigate the function of transcription tactors in induced systematic resistance primed by AR156, we detected the expression profile of 34 transcription factors in Arabidopsis treated with pathogen Pseudomonas syringae pv. tomato DC3000, after induced by AR156. We found that the expression of 17 transcription factors were affected by AR156. Since WRKY11 functions as negative regulator in SA-dependent pathway, WRKY70 functions as an important regulator in SA-ET/JA mutual antagonism and WRKY70 was up regulated and WRKY11 was down regulated by AR156, we determined studying the co-regulation of them in induced systematic resistance signal pathway by AR156. First we cloned the promoters and the CDS of these two transcription factors'genes, and than constrated the plant expression vectors WRKY11 promoter::GUS, WRKY70 promoter::GUS, CaMV 35S::WRKY11 CDS and pGreen0029::CaMV 35S promoter::WRKY70 CDS by using the unique restriction enzyme cutting sites in fragments and plant expression vectors PBI121 and pGreen 0029. After beening transformated into Escherichia coli DH5a, and correctly sequenced, they were keeped to transformate into Agrobacterium tumefaciens for future functional studies.4. ConclusionIn this study we studied the mechanism of induced systemic tolerance against drought stress by PGPR AR156 and PGPR combination, by physiological assay and molecular detection, hoping to interpret the mechanism of induced systemic drought tolerance by PGPR and be helpful for practical application. Besides, we screened transcription factors that regarded as regulators in two signal pathways stimulated during induced resistance by PGPR Bacillus cereus AR156 and constructed recombinant vectors for future functional studies of these two transcription factors.