論文 - 詳細
| RRC ID | 17437 |
|---|---|
| 著者 | Nariya H, Miyata S, Suzuki M, Tamai E, Okabe A. |
| タイトル | Development and application of a method for counterselectable in-frame deletion in Clostridium perfringens. |
| ジャーナル | Appl Environ Microbiol |
| Abstract |
Many pathogenic clostridial species produce toxins and enzymes. To facilitate genome-wide identification of virulence factors and biotechnological application of their useful products, we have developed a markerless in-frame deletion method for Clostridium perfringens which allows efficient counterselection and multiple-gene disruption. The system comprises a galKT gene disruptant and a suicide galK plasmid into which two fragments of a target gene for in-frame deletion are cloned. The system was shown to be accurate and simple by using it to disrupt the alpha-toxin gene of the organism. It was also used to construct of two different virulence-attenuated strains, ΗΝ1303 and HN1314: the former is a disruptant of the virRS operon, which regulates the expression of virulence factors, and the latter is a disruptant of the six genes encoding the α, θ, and κ toxins; a clostripain-like protease; a 190-kDa secretory protein; and a putative cell wall lytic endopeptidase. Comparison of the two disruptants in terms of growth ability and the background levels of secreted proteins showed that HN1314 is more useful than ΗΝ1303 as a host for the large-scale production of recombinant proteins. |
| 巻・号 | 77(4) |
| ページ | 1375-82 |
| 公開日 | 2011-2-1 |
| DOI | 10.1128/AEM.01572-10 |
| PII | AEM.01572-10 |
| PMID | 21183644 |
| PMC | PMC3067250 |
| MeSH | Bacterial Proteins / biosynthesis Bacterial Proteins / genetics Bacterial Toxins / biosynthesis Bacterial Toxins / genetics* Base Sequence Blotting, Northern Calcium-Binding Proteins / biosynthesis Calcium-Binding Proteins / genetics* Clostridium perfringens / enzymology Clostridium perfringens / genetics* Clostridium perfringens / metabolism Clostridium perfringens / pathogenicity Electrophoresis, Polyacrylamide Gel Hemolysin Proteins / biosynthesis Hemolysin Proteins / genetics Microbial Collagenase / biosynthesis Microbial Collagenase / genetics Mutagenesis Plasmids Reading Frames Recombinant Proteins / biosynthesis Recombinant Proteins / metabolism Sequence Analysis, Protein Sequence Deletion* Type C Phospholipases / biosynthesis Type C Phospholipases / genetics* Virulence / genetics Virulence Factors / genetics* Virulence Factors / metabolism |
| IF | 4.016 |
| 引用数 | 35 |
| WOS 分野 | BIOTECHNOLOGY & APPLIED MICROBIOLOGY MICROBIOLOGY |
| リソース情報 | |
| 一般微生物 | JCM 1419 |