Development of replicative <Emphasis Type="Italic">oriC</Emphasis> plasmids and their versatile use in genetic manipulation of <Emphasis Type="Italic">Cytophaga hutchinsonii</Emphasis> |
| |
Authors: | Yuanxi Xu Xiaofei Ji Ning Chen Pengwei Li Weifeng Liu Xuemei Lu |
| |
Institution: | (1) State Key Laboratory of Microbial Technology, College of Life Science, Shandong University, Jinan, 250100, China; |
| |
Abstract: | Cytophaga hutchinsonii is a Gram-negative aerobic soil bacterium which can digest crystalline cellulose completely through a strategy different
from that of the well-studied cellulolytic aerobic fungi and anaerobic bacteria. However, despite the availability of a published
genome sequence, studies on this organism have been very limited because of the lack of a genetic manipulation system. This
paper describes the development of replicative oriC plasmids, carrying the replication origin of the C. hutchinsonii chromosome, and an electroporation method for Escherichia coli–C. hutchinsonii shuttle vectors based on oriC plasmids with an efficiency of about 2 × 104 transformants per microgram plasmid DNA. Heterologous proteins, including green fluorescent protein and β-galactosidase,
were expressed successfully and proved functional in C. hutchinsonii under the control of the CHU_1284 promoter in oriC plasmids. Finally, the gene CHU_0344, encoding the main extracellular protein, was targeted by homologous recombination based on the oriC plasmid. These genetic techniques and tools will provide a means to study the novel cellulose degradation system of C. hutchinsonii. |
| |
Keywords: | |
本文献已被 PubMed SpringerLink 等数据库收录! |
|