Dorella F A, Estevam E M, Cardoso P G, Savassi B M, Oliveira S C, Azevedo V, Miyoshi A
Laboratório de Genética Celular e Molecular, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Vet Microbiol. 2006 May 31;114(3-4):298-303. doi: 10.1016/j.vetmic.2005.12.010. Epub 2006 Jan 26.
We developed an improved protocol for the electrotransformation of Corynebacterium pseudotuberculosis, testing variations of parameters in the procedures that are routinely used for the preparation of electrocompetent cells of this species, including (i) culture conditions, (ii) cell growth phase, (iii) electroporation solutions and (iv) quantity of plasmid DNA. We obtained the greatest efficiency of transformation when the cells were grown until the stationary phase and then washed with 10% glycerol electroporation solution. The transformation efficiency was inversely proportional to the quantity of plasmid DNA. The transformation efficiency reached 10(5) colony-forming units (cfu)/mug plasmid DNA. This protocol would be useful for genetic studies of C. pseudotuberculosis.
我们开发了一种改进的伪结核棒状杆菌电转化方案，测试了该物种感受态细胞制备常规程序中参数的变化，包括（i）培养条件、（ii）细胞生长阶段、（iii）电穿孔溶液和（iv）质粒DNA的量。当细胞生长至稳定期，然后用10%甘油电穿孔溶液洗涤时，我们获得了最高的转化效率。转化效率与质粒DNA的量成反比。转化效率达到10(5) 菌落形成单位（cfu）/μg质粒DNA。该方案将有助于伪结核棒状杆菌的遗传学研究。
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