Genetic Analysis of Flavobacterium johnsoniae (Cytophaga johnsonae) Gliding Motility

麦克布莱德 9727825滑动运动（细胞在表面上的主动运动，而不需要鞭毛的帮助）是许多不相关细菌共有的特征，但滑动运动的机制仍然是一个谜超过100年。 新开发的遗传技术将用于分析黄杆菌约翰逊（原噬纤维菌约翰逊）的滑行运动。 转座子诱变和粘粒互补将用于鉴定F。约翰逊氏菌滑行运动基因（GLD基因）。 gld基因将在E.大肠杆菌中，测序和分析。 这些序列将与序列数据库进行比较，以确定滑翔机各个部件的可能功能。 将在单个gld基因中产生突变，并评估对细胞运动性的影响。 gld基因也将被克隆到E.大肠杆菌，以允许过量生产Gld蛋白。 这些蛋白质将用于旨在确定其功能的生化实验。 它们也将被用于提高抗体。 针对单个GLD蛋白的抗体将用于测定运动细胞中每种GLD蛋白的量。 抗体还将用于通过细胞分级分离实验和通过免疫电子显微镜定位细胞内的蛋白质。 这些研究将导致一个模型的发展，以解释细菌滑行运动的机制。 除了扩大我们对这种常见但知之甚少的细菌运动方式的了解外，这些研究可能具有更大的实用价值，因为滑行细菌作为植物，动物和人类的病原体，作为生物降解的代理人，以及作为酶，抗体和其他次级代谢产物的生产者。

McBride 9727825 Gliding motility (the active movement of cells over surfaces without the aid of flagella) is a trait shared by many unrelated bacteria, yet the mechanism of gliding motility has remained a mystery for over 100 years. Newly developed genetic techniques will be used to analyze Flavobacterium johnsoniae (formerly Cytophaga johnsonae) gliding motility. Transposon mutagenesis and cosmid complementation will be used to identify F. johnsoniae gliding motility genes (gld genes). The gld genes will be cloned in E. coli, sequenced, and analyzed. The sequences will be compared to sequence databases to determine possible functions for the individual components of the gliding machinery. Mutations will created in individual gld genes and the effects on cell motility will be assessed. gld genes will also be cloned onto expression vectors in E. coli to allow overproduction of the Gld proteins. These proteins will be used for biochemical experiments aimed at determining their functions. They will also be used to raise antibodies. The antibodies against the individual GLD proteins will be used to determine the amounts of each GLD protein in the motile cells. The antibodies will also be used to localize the proteins within the cells by cell fractionation experiments and by immunoelectron microscopy. These studies will lead to the development of a model to explain the mechanism of bacterial gliding motility. In addition to expanding our knowledge on this common but little understood means of bacterial locomotion, these studies could have more practical value since gliding bacteria are important as pathogens of plants, animals, and humans, as agents of biodegredation, and as producers of enzymes, antibodies and other secondary metabolites.