| Summary: | 碩士 === 國立嘉義大學 === 生物科技研究所 === 93 === A significantly environmental and industrial contaminant, cadmium has been found in the soil polluted by wastewater from the process such as smelting of the zinc, cooper and lead, founding of the coating, lead-acid storage batteries, alloy, paint and plastic. In previous studies, several cadmium-resistant microorganism have been isolated from these polluted environment. The mechanism resistant to cadmium include 1) active transport of the metal away from the microorganism, 2) metal exclusion by permeability, 3) intracellular sequestration of metals by protein binding, 4) extracellular sequestration, and 5) reduction metal sensitivity of cellular targets. Here, cadmium-polluted farms have been detected Taoyuan, Yunlin, and Tainan. To eliminate such contamination, the microorganism capable of depolluting cadmium from environment would be the best choice. Therefore, the purpose of this study was to discover the cadmium-resistant microorganisms and then to determine their resistance mechanism. 20 cadmium-resistant organisms were isolated from cadmium-polluted and tainted soil by a waterink factory in Yunlin and then cadmium tolerance was separated into two groups: lower than 400 ppm and ranges from 650 to 900 ppm, respectively. Some organisms were tested by BIOLOG bacterial identification system and identified these organisms, Isolates rcd1, 6, 7, 17, 18 and 20 appeared to be Ralstonia eutrophus. confirmation of These organism were furthern confirmed with molecular method by nucleotide sequence of 16S rDNA. The results suggested that strains rcd 12, 14and 21 were Ralstonia taiwanensis strain LMG19425-like organism; strains strains rcd 2, 3, 4, 8, and 9 were belonged to genus Ralstonia; the remaining strains rcd 5, 11, 13, 15, and 16 were related to Streptomyces spp. Using czc operon, a efflux pump for cadmium resistance as PCR target, all Ralstonia strains consisted of this operon. To elucidate this phenomenon, plasmid transfer might be responsible. Plasmid profile was constructed by alkaline method and showed 8 of Ralstonia strains, rcd 2, 3, 4, 7, 17, 18, 19,and 20, harbored a large plasmid with the size over 150 kb. Whether czc operon was located on chromosome or plasmid, Southern blot analysis was performed by using czcA as probe. The results revealed that this operon was located on plasmid and chromosome as well. Furthermore, we will determine the conjugation ability of these plasmids. Here we first reported that cadmium-resistant Ralstonia strains use czc operon, which is located on plasmid and chromosome, to release cadmium off the organism.
|
|---|
