Abstract
Bukholderia glumae is a gram-negative bacterium which causes grain rot, seedling rot and panicle blight in rice and bacterial wilt in many field crops. This bacterium has been reported from major rice growing regions around the world and is now considered as an emerging major pathogen of rice (Tsushima et al., 1996; Jeong et al., 2003; Kim et al., 2010; Ham et al., 2011). Here we describe two methods for competent cells preparation and transformation of B. glumae. Using these methods, we have applied effector detector system (Sohn et al., 2007) to B. glumae (Sharma et al., 2013).
Keywords: Burkholderia glumae, Competent cells, Transformation, PEDV5 based vectors
Materials and Reagents
Equipment
Procedure
Part I: Conventional method
Part II. High competency method
Recipes
Acknowledgments
This protocol was adapted from Sharma et al. (2013). This work was supported by the ‘Program for Promotion of Basic Research Activities for Innovative Biosciences (PROBRAIN)’ (Japan) and Japan Society for the Promotion of Science (JSPS) grants no. 18310136 and 20380027, and ‘The Ministry of Agriculture, Forestry, and Fisheries of Japan (Genomics for Agricultural Innovation PMI-0010)’ and the Ministry of Education, Culture, Sports, Science and Technology of Japan (Grant-in-Aid for Scientific Research on Innovative Areas 23113009); and by JSPS grant nos 22780040 and 2301518 to H. Saitoh, JSPS grant no. 2200214 to R. Terauchi. The financial assistance received from JSPS to Shailendra Sharma and Shiveta Sharma for carrying out this study is gratefully acknowledged. S. Kamoun and J.D.G. Jones were supported by The Gatsby Foundation (United Kingdom).
References
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We haven't used 2mm cuvette for high competency method. But you can try it according to the setting written in conventional method.
I couldn't replicate the method using my Burkholderia isolate. The electroporator says arc detected, meanwhile the conventional method works fine. I wonder what's the possibility of failure during pulsing..