https://scholars.lib.ntu.edu.tw/handle/123456789/96215
Title: | 桿菌屬細菌表現系統之建立及應用─桿菌屬拮抗細菌之開發與改良(3/3) | Authors: | 陳昭瑩 | Issue Date: | 1999 | Publisher: | 臺北市:國立臺灣大學植物病理與微生物學系暨研究所 | Abstract: | 已知許多細菌可產生幾丁質分解 酵素分解真菌細胞壁,抑制真菌的生 長,故被應用於植物真菌性病害的防 治上。本研究將Bacillus circulans chiA 基因導入拮抗細菌Bacillus subtilis 細 胞中,測試其對拮抗菌抑制病原真菌 生長的影響。首先將含起動子的chiA 基因次選殖於shuttle vector pHY300PLK 上,以原生質體轉形法送 入B. subtilis 細胞中。可於B. subtilis 的培養上清液中測得幾丁質分解酵素 的活性,顯示ChiA1 蛋白質可分泌於 B. subtilis 的細胞外。B. subtilis 於LB 中培養24 小時後可以測得幾丁質分解 酵素的活性;在幾丁質培養基中則需 培養較長時間才可測得幾丁質分解酵 素的活性。在大腸桿菌細胞中,ChiA1 蛋白質主要分泌於細胞間質中。當培 養時間延長時,可於幾丁質培養基上 菌落外緣出現透明圈。將B. circulans chiA 基因表現於無抑菌作用的變異 株, 可以觀察到ChiA 對Botrytis elliptica 的生長具有抑制效果;但幾丁 質分解酵素ChiA1 並不能穩定地促進 B. subtilis 野生菌株的抑菌效果。 Several chitinase - producing bacteria are known to degrade the fungal cell wall and are able to inhibit the growth of fungi. Applications of chitinase-producing bacteria in plant disease control have been reported. In this study, we transferred the chiA gene of chitinase-producing bacterium, Bacillus circulans, into the antagonist, Bacillus subtilis and examined the effect of ChiA1 protein on the antagonistic activity of B. subtilis. Firstly, the chiA gene was subcloned with its own promoter into the shuttle vector, pHY300PLK, and transferred into the bacterial cells of Bacillus subtilis by protoplast transformation. The expression of ChiA1 protein was detected in the supernatant of B. subtilis culture, indicating that the ChiA1 2 protein is secreted out of the Bacillus cell. The ChiA1 protein was detected 24 hr after cultivation of Bacillus in LB broth and in a later period when incubated in chitin medium. In Escherichia coli cells, the ChiA1 protein is translocated into the periplasmic space. The chitinase is released from E. coli cells after prolonged incubation. The expression of chiA gene in a non-antagonistic mutant of B. subtilis could display inhibition effect on the growth of Botrytis elliptica . However, when the chiA gene was expressed in the antagonistic strain of B. subtilis, the enhancing effect of ChiA1 on the antagonistic activity of B. subtilis was not readily observed. |
URI: | http://ntur.lib.ntu.edu.tw//handle/246246/29752 | Other Identifiers: | 882316B002016B19 | Rights: | 國立臺灣大學植物病理與微生物學系暨研究所 |
Appears in Collections: | 植物病理與微生物學系 |
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882316B002016B19.pdf | 45.88 kB | Adobe PDF | View/Open |
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