| 研究生: |
簡勤益 Chin-I Chien |
|---|---|
| 論文名稱: |
Galectin-1枯草桿菌、阿拉伯芥與加洲苜蓿夜蛾核多角體病毒轉殖株的建立及殺蟲活性之研究 Establishment and Characterization of Insecticidal Galectin-1 Transgenic Strains of Bacillus subtilis, Arabidopsis thaliana and Autographa californica Multiple Nucleopolyhedrovirus |
| 指導教授: |
黃榮南
Rong-Nan Huang |
| 口試委員: | |
| 學位類別: |
碩士 Master |
| 系所名稱: |
生醫理工學院 - 生命科學系 Department of Life Science |
| 畢業學年度: | 95 |
| 語文別: | 英文 |
| 論文頁數: | 64 |
| 中文關鍵詞: | 枯草桿菌 、加洲苜蓿夜蛾核多角體病毒 、阿拉伯芥 、生物殺蟲劑 |
| 外文關鍵詞: | Autographa californica Multiple Nucleopolyhedrov, bio-insecticide, Bacillus subtilis, Galectin-1, Arabidopsis thaliana |
| 相關次數: | 點閱:8 下載:0 |
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先前,我們實驗室的研究結果已指出,Galectin-1 (GAL1) 是一個幾丁質結合蛋白,而且對小菜蛾具有殺蟲活性。在本篇研究中,將倉鼠卵巢細胞的GAL1基因序列,建構在pαHY300、pCAMBIA1390-35S和pBac-IR-GFP的表現載體上,並將載體分別命名為pαHY300-GAL1、p1390-35S-GAL和pBac-GAL1-IR-GFP,並且將他們分別送入枯草桿菌、阿拉伯芥和加洲苜蓿夜蛾核多角體病毒中,製成GAL1轉殖株,進行GAL1蛋白質的表現及其殺蟲活性的研究。而結果指出GAL1基因確實成功地送入枯草桿菌、阿拉伯芥和核多角體病毒中,並且都會表現GAL1蛋白質,在GAL1蛋白質表現量方面,阿拉伯芥和核多角體病毒遠高於枯草桿菌,在殺蟲活性的結果中,枯草桿菌GAL1轉殖株對小菜蛾及斜紋夜盜沒有明顯的殺蟲活性,而阿拉伯芥GAL1轉殖株對小菜蛾及斜紋夜盜則有明顯的殺蟲活性,核多角體病毒GAL1轉殖株的殺蟲活性,仍在進行當中。而以上的結果指出,以GAL1蛋白質做為新型的具有殺蟲活性的轉殖植物是一個很好的選擇。
Previous studies in our laboratory have indicated that galectin-1 (GAL1) is a chitin binding protein and shows insecticidal activity toward Plutella xylostella. In these studies, GAL1 gene from Chinese hamster ovary cells was cloned into the plasmids pαHY300, pCAMBIA1390-35S and pBac-IR-GFP respectively. The resulting plasmids were designed as pαHY300-GAL1, p1390-35s-GAL1 and AcMNPV-GAL1 and introduced into Bacillus subtilis, Arabidopsis thaliana and Autographa californica multiple nucleopolyhedrovirus respectively for GAL1 expression and insecticidal activity investigation. The results showed that GAL1 gene was successfully introduced into B. subtilis, A. thaliana and AcMNPV for protein expression. The expression levels of GAL1 were highest in A. thaliana among the transformants. Though the GAL1-transgenic B. subtilis showed no significant insecticidal activity to S. litura and P. xylostella, GAL1-transgenic A. thaliana showed high insecticidal activity toward Spodoptera litura and Plutella xylostella. The insecticidal activities of GAL1-transfected AcMNPV will be under investigation. The results indicate that GAL1 would be a future candidate gene in transgenic plants.
Ahmed H., Pohl J., Fink N. E. , Strobel F. and Vasta G. R. (1996). The primary structure and carbohydrate specificity of a beta-galactosyl-binding lectin from toad (Bufo arenarum Hensel) ovary reveal closer similarities to the mammalian galectin-1 than to the galectin from the clawed frog Xenopus laevis. J. Biol. Chem. 271:33083-94.
Bianchet M. A., Ahmed H., Vasta G. R. and Amzel L. M. (2000). Soluble beta-galactosyl-binding lectin (galectin) from toad ovary: crystallographic studies of two protein-sugar complexes. Proteins. 40:378-88.
Boldt D. H. and Banwell J. G. (1985). Binding of isolectins from red kidney bean (Phaseolus vulgaris) to purified rat brush-border membranes. Biochim. Biophys. Acta. 843:230-7.
Briles E. B., Gregory W, Fletcher P. and Kornfeld S. (1979). Vertebrate lectins, Comparison of properties of beta-galactoside-binding lectins from tissues of calf and chicken. J. Cell Biol. 81:528-37.
Brockmeier U., Wendorff M. and Eggert T. (2006). Versatile expression and secretion vectors for Bacillus subtilis. Curr. Microbiol. 52:143-8.
Carlini C. R. and Grossi-de-Sa M. F. (2002). Plant toxic proteins with insecticidal properties. A review on their potentialities as bioinsecticides. Toxicon. 40:1515-39.
Cerra R. F., Haywood-Reid P. L. and Barondes S. H. (1984). Endogenous mammalian lectin localized extracellularly in lung elastic fibers. J. Cell Biol. 98:1580-9.
Chang J. H., Choi J. Y., Jin B. R., Roh J. Y., Olszewski J. A., Seo S. J., O''Reilly D. R. and Je Y. H. (2003). An improved baculovirus insecticide producing occlusion bodies that contain Bacillus thuringiensis insect toxin. J. Invertebr. Pathol. 84:30-7.
Chang Y. Y., Chen S. J., Liang H. C., Sung H. W., Lin C. C. and Huang R. N. (2004). The effect of galectin 1 on 3T3 cell proliferation on chitosan membranes. Biomaterials. 25:3603-11.
Chakrabarty R., Viswakarma N., Bhat S. R., Kirti P. B., Singh B. D. and Chopra V. L. (2002). Agrobacterium-mediated transformation of cauliflower: optimization of protocol and development of Bt-transgenic cauliflower. J. Biosci. 27:495-502.
Chen Y. J., Chen W. S. and Wu T. Y. (2005). Development of a bi-cistronic baculovirus expression vector by the Rhopalosiphum padi virus 5'' internal ribosome entry site. Biochem. Biophys. Res. Commun. 335:616-23.
Chen N. T. (2005). Characterization of the insecticidal activity of galectin-1 on Plutella xylostella. 中央大學碩士論文.
Chen Y. C. (2001). Cloning and expression of Galectin-1 and Thioreoredoxin peroxidaseⅡ genes. 中央大學碩士論文.
Chrispeels M. J. and Raikhel N. V. (1991). Lectins, lectin genes, and their role in plant defense. Plant Cell. 3:1-9.
Cleves A. E., Cooper D. N., Barondes S. H. and Kelly R. B. (1996). A new pathway for protein export in Saccharomyces cerevisiae. J. Cell Biol. 133:1017-26.
Cooper D. N. and Barondes S. H. (1990). Evidence for export of a muscle lectin from cytosol to extracellular matrix and for a novel secretory mechanism. J. Cell Biol. 110:1681-91.
Cooper D. N., Massa S. M. and Barondes S. H. (1991). Endogenous muscle lectin inhibits myoblast adhesion to laminin. J. Cell Biol. 115:1437-48.
Cooper D. N. (2002). Galectinomics: finding themes in complexity. Biochim Biophys Acta. 1572:209-31.
Dodd J. and Jessell T. M. (1986). Cell surface glycoconjugates and carbohydrate-binding proteins: possible recognition signals in sensory neurone development. J. Exp. Biol. 124:225-38.
Donatucci D.A., Liener I. E. and Gross C .J. (1987). Binding of navy bean (Phaseolus vulgaris) lectin to the intestinal cells of the rat and its effect on the absorption of glucose. J. Nutr. 117:2154-60.
Endo Y., Oka T., Tsurugi K. and Franz H. (1989). The mechanism of action of the cytotoxic lectin from Phoradendron californicum: the RNA N-glycosidase activity of the protein. FEBS Lett. 248:115-8.
Fink de Cabutti N. E., Caron M., Joubert R., Elola M. T., Bladier D. and Herkovitz J. (1987). Purification and some characteristics of a beta-galactoside binding soluble lectin from amphibian ovary. FEBS Lett. 223:330-4.
Fred Brewer C. (2002). Binding and cross-linking properties of galectins. Biochim Biophys Acta. 1572:255-62.
Goldstein I. J. and Hayes C. E. (1978). The lectins: carbohydrate-binding proteins of plants and animals. Adv. Carbohydr. Chem. Biochem. 35:127-340.
M. S. Harper, T. L. Hopkins and T. H. Czapla (1998). Effect of wheat germ agglutinin on formation and structure of the peritrophic membrane in european corn borer (Ostrinia nubilalis) larvae. Tissue and Cell. 30:166-176.
Huesing J. E, Shade R. E, Chrispeels M. J. and Murdock L. L. (1991). alpha-Amylase Inhibitor, Not Phytohemagglutinin, Explains Resistance of Common Bean Seeds to Cowpea Weevil. Plant Physiol. 96:993-996.
Hynes M. A., Buck L. B., Gitt M., Barondes S., Dodd J. and Jessell T. M. (1989). Carbohydrate recognition in neuronal development: structure and expression of surface oligosaccharides and beta-galactoside-binding lectins. Ciba Found Symp. 145:189-210.
Janzen D. H., Juster H. B. and Liener I. E. (1976). Insecticidal action of the phytohemagglutinin in black beans on a bruchid beetle. Science. 192:795-6.
Kumar M. A., Timm D. E., Neet K. E., Owen W. G., Peumans W. J. and Rao A. G. (1993). Characterization of the lectin from the bulbs of Eranthis hyemalis (winter aconite) as an inhibitor of protein synthesis. J Biol Chem. 268:25176-83.
Liu F. T., Patterson R. J. and Wang J. L. (2002). Intracellular functions of galectins. Biochim Biophys Acta. 1572:263-73.
Pusztai A, Ewen S. W., Grant G., Peumans W.J., Van Damme E.J., Coates M. E., and Bardocz S. (1995). Lectins and also bacteria modify the glycosylation of gut surface receptors in the rat. Glycoconj. J. 12:22-35.
Rao K.V., Rathore K.S., Hodges T.K., Fu X., Stoger E., Sudhakar D., Williams S., Christou P., Bharathi M., Bown D. P., Powell K. S., Spence J., Gatehouse A. M. and Gatehouse J. A. (1998). Expression of snowdrop lectin (GNA) in transgenic rice plants confers resistance to rice brown planthopper. Plant J. 15:469-77.
Rini J. M. and Lobsanov Y. D. (1999). New animal lectin structures. Curr Opin Struct Biol. 9:578-84.
Sehgal D., Malik P. S. and Jameel S. (2003). Purification and diagnostic utility of a recombinant hepatitis E virus capsid protein expressed in insect larvae. Protein Expr. Purif. 27:27-34
Szewczyk B., Hoyos-Carvajal L., Paluszek M., Skrzecz I. and Lobo de Souza M. (2006). Baculoviruses-- re-emerging biopesticides. Biotechnol. Adv. 24:143-60.
Van Frankenhuyzen K, Gringorten J. L, Milne R. E, Gauthier D, Pusztai M, Brousseau R. and Masson L. (1991). Specificity of Activated CryIA Proteins from Bacillus thuringiensis subsp. kurstaki HD-1 for Defoliating Forest Lepidoptera. Appl. Environ. Microbiol. 57:1650-1655.
Wasano K., Hirakawa Y. and Yamamoto T. (1990). Immunohistochemical localization of 14 kDa beta-galactoside-binding lectin in various organs of rat. Cell Tissue Res. 259:43-9.
Zheng R. J. (2005). Cloning and expression of cry1Aa, cry1Ab, cry1C and cry1Da genes from Bacillus thuringiensis subsp. aizawai. 朝陽大學碩士論文.
Zhou Q. and Cummings R. D. (1993). L-14 lectin recognition of laminin and its promotion of in vitro cell adhesion. Arch. Biochem. Biophys. 300:6-17.