FLP-FRT-based Method to Obtain Unmarked Deletions of CHU_3237 (porU) and Large Genomic Fragments of Cytophaga Hutchinsonii

Overview

Journal Appl Environ Microbiol

Specialties Environmental Health
Microbiology

Date 2014 Jul 27

PMID 25063660

Citations 20

Authors

Ying Wang

Zhiquan Wang

Jing Cao

Zhiwei Guan

Xuemei Lu

Affiliations

Soon will be listed here.

Abstract

Cytophaga hutchinsonii is a widely distributed cellulolytic bacterium in the phylum Bacteroidetes. It can digest crystalline cellulose rapidly without free cellulases or cellulosomes. The mechanism of its cellulose utilization remains a mystery. We developed an efficient method based on a linear DNA double-crossover and FLP-FRT recombination system to obtain unmarked deletions of both single genes and large genomic fragments in C. hutchinsonii. Unmarked deletion of CHU_3237 (porU), an ortholog of the C-terminal signal peptidase of a type IX secretion system (T9SS), resulted in defects in colony spreading, cellulose degradation, and protein secretion, indicating that it is a component of the T9SS and that T9SS plays an important role in cellulose degradation by C. hutchinsonii. Furthermore, deletions of four large genomic fragments were obtained using our method, and the sizes of the excised fragments varied from 9 to 19 kb, spanning from 6 to 22 genes. The customized FLP-FRT method provides an efficient tool for more rapid progress in the cellulose degradation mechanism and other physiological aspects of C. hutchinsonii.

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