Dissertation > Excellent graduate degree dissertation topics show

Isolation and Identification of Endophytic Bacteria BEB3 for Siderophores Product from Banana and Preliminary Studies on Mechanism of Biological Control

Author: ZhangKeLi
Tutor: HuangGuiXiu
School: Hainan University
Course: Agricultural biotechnology
Keywords: endophytic bacteria Fusarium oxysporum f. sp. Cubense siderophore antagonistic effect Green Fluorescent Protein promote growth
CLC: S436.68
Type: Master's thesis
Year: 2010
Downloads: 87
Quote: 0
Read: Download Dissertation

Abstract

71 endophytic bacterial strains were isolated from banana in plantation of Tropical Cropos Genetic Resources Institute of Chinese Academy of Tropical Agricultural Sciences. The endophytic bacterial content of GCTCV106 and TMBX1378 varieties were the maximum, reached up to 1×106cfu/g. The content in Taijiao III was the minimum, also reach to 1×104 cfu/g. Six endophytic bacterial isolates (8.45% of total) were antagonistic to Fusarium oxysporum f. sp. Cubense(FOC4) by in vitro evaluation. The antagonistic effect of isolate BEB3 was the best. Through morphological, physiological, biochemical and 16S rDNA molecular identification, BEB3 was determined to Raul’s genus(Ralstonia sp.), and the homologous similarity compared with Ralstonia mannitolilytica was up to 99.6%.The mutant strains of BEB3 antagnonisted 200μg/ml rifampicin were screened by Gradient induction. The suicide plasmid pJBA28 with GFP gene was mobilized into BEB3 strains by conjugation. The positive clones grown on the dual-resistant plate were selected and subsequently identified by PCR, fluorescence confocal microscopy and southern hybridization. Banana plantlets were inoculated with root soaking method. The DNA of banana seedlings were extracted after inoculated 10 d,20 d,30 d. BEB3 strains tagged by GFP was tested with PCR. The result showed that BEB3 could colonize within the banana plant.The tasting results revealed that BEB3 could produce siderophore in CAS agar Plate. Fermentation conditions were optimized. Single factor experiment and orthogonal experiments showed that the optimum composition for the fermentation medium of its growth quantity (1 L) was:glucose 4 g, cascamino acid 4 g,1 mol/L MgCl2 0.5 ml,1 mol/L CaCl2 1 ml. However the optimum composition for the fermentation medium of production of siderophore (1L) was:glucose 1 g, cascamino acid 1 g,1 mol/L MgCl2 0.5 ml,1 mol/L CaCl2 1 ml. Consideration of fementation cost、the bacteria’s growth quantity and its production of siderophore showed that the optimum fermentation conditions were:temperature 28℃, media amount 50 mL/150 mL, original pH6.5, inouclation amount7%, the concentration of Fe3+0.4 mg/L.The preliminary research of resistance evaluation of BEB3 showed that its germfree filtrate can inhibite spore germination and mycelial growth of the Fusarium oxysporum. The germfree filtrate had high thermal stability. Folllowed in 100℃for 10 min, the antibacterial effect had no change. While in 121℃for 10 min, the antibacterial effect still reached to 58.2%.with three different ways to inoculate the banana’s leaves in vitro, the results shows that the leaves have no disease signs in the way that inoculate pathogen after inoculate BEB3 2 days.from pot experiment,the incidence rate of banana with endophytic bacteria is 8.33%,but the control is 85.71%.the endophytic bacteria could promote the banana growth in sand bed seedlings and in pot seedlings.

Related Dissertations

CLC: > Agricultural Sciences > Plant Protection > Pest and Disease Control > Horticultural Crops Pest and Disease Control > Fruit tree pests and diseases > Perennial herbaceous fruit pests and diseases
© 2012 www.DissertationTopic.Net Mobile