کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10889816 | 1081500 | 2010 | 9 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Application of an inducible system to engineer unmarked conditional mutants of essential genes of Pseudomonas aeruginosa
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موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
بیوتکنولوژی یا زیستفناوری
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چکیده انگلیسی
The ΦCTX-based integration vector pYM101 harboring a tightly controlled modified phage T7 early gene promoter/LacIq repressor (T7/LacI) system was constructed for the generation of unmarked conditional mutants in Pseudomonas aeruginosa. Promoter activity of the T7/LacI system was demonstrated to be dependent on the presence of the inducer isopropyl -β-D-1-thiogalactopyranoside (IPTG), as evaluated by measuring β-galactosidase activity. In the absence of the inducer, the promoter was silent as its activity was lower than those of a promoter-less lacZ control. Unmarked conditional mutants of four predicted essential genes (lolCDE (PA2988-86), lpxC (PA4406), rho (PA5239), and def (PA0019)) were successfully constructed using this recombination system. In the absence of IPTG, the growth of all mutants was repressed; however, the addition of either 0.1 or 1 mM IPTG restored growth rates to levels nearly identical to wild-type cells. It was therefore demonstrated that the inducible integration vector pYM101 is suitable for the creation of unmarked conditional mutants of P. aeruginosa, and is particularly useful for examining the function of essential genes.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Microbiological Methods - Volume 82, Issue 3, September 2010, Pages 205-213
Journal: Journal of Microbiological Methods - Volume 82, Issue 3, September 2010, Pages 205-213
نویسندگان
Yuji Morita, Shin-Ichiro Narita, Junko Tomida, Hajime Tokuda, Yoshiaki Kawamura,