Burkholderia glumae Competent Cells Preparation and Transformation
水稻细菌性谷枯病菌感受态细胞的制备和转化
发布: 2013年12月05日第3卷第23期 DOI: 10.21769/BioProtoc.985 浏览次数: 11698
评审: Tie Liu
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参见作者原研究论文
The authors used this protocol in:
May 2013
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 (水稻细菌性谷枯病菌)
Competent cells (感受态细胞)
Transformation (转型)
PEDV5 based vectors (pedv5载体)
Materials and Reagents
- Burkholderia glumae strain 106619 (National Institute of Agricultural Sciences Genebank, Tsukuba, Ibaraki, Japan)
- pEDV5 based vectors (Fabro et al., 2011; Sharma et al., 2013)
- Gentamycin
- Tryptone
- Yeast extract
- Protease peptone no. 3
- Ice
- Lysogeny Broth (LB) medium (see Recipes)
- King’s Broth (KB) agar (see Recipes)
- KB agar with 25 ng/ml gentamycin (see Recipes)
- 10% glycerol (see Recipes)
- 300 mM sucrose solution (see Recipes)
Equipment
- Deep freezer
- Autoclave
- Clean bench
- Petri plates
- Sterile 1.5 ml tubes
- Sterile 15 ml tubes
- Sterile 50 ml tubes
- Incubation Shaker
- Aluminum foil
- Spectrophotometer
- Centrifuge
- Toothpick
- MicroPulser/Gene Pulser Cuvettes, 0.2 cm gap (Bio-Rad Laboratories, catalog number: 165-2089 )
- MicroPulser/Gene Pulser Cuvettes, 0.1 cm gap (Bio-Rad Laboratories, catalog number: 165-2086 )
- Gene Pulser XcellTM Electroporation Systems (Bio-Rad Laboratories)
- Minisart filters (pore size 0.2 μm) (Sigma-Aldrich, catalog number: 16534K )
- Disposable Cell Spreaders
Procedure
文章信息
版权信息
© 2013 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Saitoh, H. and Terauchi, R. (2013). Burkholderia glumae Competent Cells Preparation and Transformation. Bio-protocol 3(23): e985. DOI: 10.21769/BioProtoc.985.
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分类
微生物学 > 微生物-宿主相互作用 > 体内实验模型 > 植物
植物科学 > 植物免疫 > 信号感知与传递
分子生物学 > DNA > 转化
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