Universität Bonn, Institut für Mikrobiologie & Biotechnologie, 168 Meckenheimer Allee, 53515 Bonn, Germany.
J Biotechnol. 2010 Dec;150(4):460-5. doi: 10.1016/j.jbiotec.2010.10.069. Epub 2010 Oct 20.
The characteristic ability of Gluconobacter oxydans to incompletely oxidize numerous sugars, sugar acids, polyols, and alcohols has been exploited in several biotechnological processes, for example vitamin C production. The genome sequence of G. oxydans 621H is known but molecular tools are needed for the characterization of putative proteins and for the improvement of industrial strains by heterologous and homologous gene expression. To this end, promoter regions for the genes encoding G. oxydans ribosomal proteins L35 and L13 were introduced into the broad-host-range plasmid pBBR1MCS-2 to construct two new expression vectors for gene expression in Gluconobacter spp. These vectors were named pBBR1p264 and pBBR1p452, respectively, and have many advantages over current vectors for Gluconobacter spp. The uidA gene encoding β-D-glucuronidase was inserted downstream of the promoter regions and these promoter-reporter fusions were used to assess relative promoter strength. The constructs displayed distinct promoter strengths and strong (pBBR1p264), moderate (pBBR1p452) and weak (pBBR1MCS-2 carrying the intrinsic lac promoter) promoters were identified.
氧化葡萄糖杆菌（Gluconobacter oxydans）不完全氧化许多糖、糖酸、多元醇和醇的特征能力已在几种生物技术过程中得到利用，例如维生素 C 的生产。氧化葡萄糖杆菌 621H 的基因组序列是已知的，但需要分子工具来表征假定的蛋白质，并通过异源和同源基因表达来改良工业菌株。为此，将编码氧化葡萄糖杆菌核糖体蛋白 L35 和 L13 的基因的启动子区域引入到广谱宿主范围质粒 pBBR1MCS-2 中，以构建两种用于在葡萄糖杆菌属中表达基因的新表达载体。这些载体分别命名为 pBBR1p264 和 pBBR1p452，与当前用于葡萄糖杆菌属的载体相比具有许多优势。将编码β-D-葡糖醛酸酶的 uidA 基因插入启动子区域的下游，并使用这些启动子-报告基因融合来评估相对启动子强度。这些构建体显示出不同的启动子强度，确定了强（pBBR1p264）、中（pBBR1p452）和弱（携带固有 lac 启动子的 pBBR1MCS-2）启动子。
