کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8422379 | 1545945 | 2014 | 6 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Optimization of a plasmid electroporation protocol for Aeromonas salmonicida subsp. salmonicida
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
بیوتکنولوژی یا زیستفناوری
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چکیده انگلیسی
Aeromonas salmonicida subsp. salmonicida is a major fish pathogen. Molecular tools are required to study the virulence and genomic stability of this bacterium. An efficient electroporation-mediated transformation protocol for A. salmonicida subsp. salmonicida would make genetic studies faster and easier. In the present study, we designed the 4.1-kb pSDD1 plasmid as a tool for optimizing an electroporation protocol for A. salmonicida subsp. salmonicida. We systematically tested the electroporation conditions to develop a protocol that generates the maximum number of transformants. Under these optimal conditions (25 kV/cm, 200 Ω, 25 μF), we achieved an electroporation efficiency of up to 1 Ã 105 CFU/μg DNA. The electroporation protocol was also tested using another plasmid of 10.6-kb and three different strains of A. salmonicida subsp. salmonicida. The strains displayed significant differences in their electro-transformation competencies. Strain 01-B526 was the easiest to electroporate, especially with the pSDD1 plasmid. This plasmid was stably maintained in the 01-B526 transformants, as were the native plasmids, but could be easily cured by removing the selection conditions. This is the first efficient electroporation protocol reported for A. salmonicida subsp. salmonicida, and offers new possibilities for studying this bacterium.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Microbiological Methods - Volume 98, March 2014, Pages 44-49
Journal: Journal of Microbiological Methods - Volume 98, March 2014, Pages 44-49
نویسندگان
Stéphanie Dallaire-Dufresne, Jean-Guillaume Emond-Rheault, Sabrina A. Attéré, Katherine H. Tanaka, Mélanie V. Trudel, Michel Frenette, Steve J. Charette,