Studies of low-molecular-weight-bacteriocin Carocin protein secretion mechanism in Pectobacterium carotovorum subsp. carotovorum

Autor: Ying-chih Lo, 羅英誌
Rok vydání: 2013
Druh dokumentu: 學位論文 ; thesis
Popis: 101
Pectobacterium carotovorum subsp. carotovorum (Pcc) is a Gram-negative enterobacterium that is known to cause soft-rot disease on plants including economically crops such as Chinese cabbage, celery, potato, and orchid. When it was subjected to adverse circumstances such as high population density, nutrient depletion and UV light, it would secrete one or more antibacterial substances called bacteriocins. Bacteriocins which are antimicrobial and toxic proteins inhibit growth of the related bacteria strains. There is no doubt that bacteriocins produced from Pcc not only provides a biological method for soft-rot disease of plants but also not easily causes environmental pollution. In this studies, the genes that are related to bacteriocin production is a pivotal factor to solve problem about soft-rot disease of many economical crops. From previous studies, low-molecular-weight bacteriocin Carocin S1 had already been confirmed and reported that it was exported through the bacterial flagellum type III secretion system. The bacterial flagellum secretion system consisting of more than 20 different proteins is a rotary device for motility. Interestingly, The sequence of the gene related to LMWB is homologous to proteins of the type III secretion system injectisome. It seems that LMWB produced from Pcc not simply contribute to the bacterial flagellum type III secretion system. In this study, flagellin gene (fliG) of Pcc (H-rif-8-6 was used in this study) was separated by Polymerase chain reaction (PCR) and cloned into pET32a expression vector. The recombinant flagellin was purified by affinity purification using Ni2+ -Sepharose resin. Furthermore, the mouse and New Zealand White rabbit were immunized with both flagellin FliC and FliG respectively to obtain polyclonal antiserum. Western blot analysis will be elaborated in the following paragraphs about results. The interaction between FliC and FliG protein are investigated by immunoprecipitation experiment. The results reveal they did not form complex and suggest possible LMWB secretion mechanism.
Databáze: Networked Digital Library of Theses & Dissertations
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