Quantitative Evaluation of Competitive Nodulation among Different Sinorhizobium Strains
不同中华根瘤菌菌株中竞争性结瘤的量化评估
发布: 2015年08月05日第5卷第15期 DOI: 10.21769/BioProtoc.1555 浏览次数: 8095
评审: Zhaohui LiuAnonymous reviewer(s)
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参见作者原研究论文
The authors used this protocol in:
Apr 2014
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Abstract
Legumes play a vital role in global food supply because they are uniquely capable of fixing atmospheric nitrogen (N) through symbioses with root and stem nodule bacteria, collectively called the rhizobia. These commonly include bacteria in the genera Rhizobium, Mesorhizobium, Sinorhizobium (Ensifer), and Bradyrhizobium, although other genera of bacteria have now been shown to form root nodule symbioses with several legume species (Weir, 2012). The symbiotic interaction is important for agricultural productivity, especially in less developed countries where nitrogen fertilizer is expensive. However, nodulation ability and competitiveness have practical importance in agricultural production, because the inoculation of efficient rhizobia is often unsuccessful, due to large part to the presence of competitive populations of ineffective indigenous rhizobia in soils (Toro, 1996; Triplett and Sadowsky, 1992). This protocol allows one us to quantitatively evaluate the relative nodulation competitiveness of Sinorhizobium strains.
Materials and Reagents
- Two Sinorhizobium meliloti (S. medicae) strains (strain A and B) with different intrinsic antibiotic resistance (e.g. Strain A is neomycin sensitive and strain B is neomycin resistant at a certain concentration of neomycin)
Note: Alternately, strains can be differentiated by using gfp and rfp, antisera, or other marker genes (Triplett and Sadowsky, 1992).
- Medicago truncatula seeds
- Sodium hypochlorite
- Sodium chloride (Macron Chemical, catalog number: 7581-06 )
- Concentrated sulfuric acid (Thermo Fisher Scientific, catalog number: A300-212 )
- Ethanol (Decon Labs, catalog number: 2701 )
- Antibiotics
- Sunshine mix #5:Turface mixture at 1: 1 ratio (SunGro Horticulture)
- Turface MVP (Profile Product LLC)
- Tryptone (BD, catalog number: 211699 )
- Yeast extract (Thermo Fisher Scientific, catalog number: DF0127071 )
- CaCl2.2H2O (Thermo Fisher Scientific, catalog number: C79-3 )
- Agar (Sigma-Aldrich, catalog number: A1296 )
- KNO3 (Thermo Fisher Scientific, catalog number: P263-500 )
- Ca(NO3)2.4H2O (Acros, catalog number: 423535000 )
- Ca(H2PO4)2 (Spectrum, catalog number: C1145 )
- MgSO4.7H2O (J.T.Baker®, catalog number: 2504 )
- Fe-EDTA (Sigma-Aldrich, catalog number: E6760-5000 )
- MnCl2 (Thermo Fisher Scientific, catalog number: M33-500 )
- H3BO3 (Mallinckrodt, catalog number: 2549 )
- ZnSO4.7H2O (J.T.Baker®, catalog number: 4382 )
- NaMoO4 (Sigma-Aldrich, catalog number: S6646 )
- CuSO4.5H2O (Thermo Fisher Scientific, catalog number: C-493 )
- K2SO4 (EMD Millipore, catalog number: PX1595-1 )
- CaSO4.2H2O (Sigma-Aldrich, catalog number: C3771 )
- Paraffin (Thermo Fisher Scientific, catalog number: P-21 hard )
- TY medium (see Recipes)
- N2-free nutrient solution (see Recipes)
- Sterile paraffin coated sands (see Recipes)
Equipment
- MAGENTA® vessel GA-7 (Sigma-Aldrich, catalog number: V8505 ) and cotton rope (diameter ¼ inch) for Leonard jar assemblies
- Filter paper P5 (Thermo Fisher Scientific, catalog number: 09-801C )
- Plant growth chamber (TC2, Environmental Growth Chambers as an example but other plant growth chambers can be used)
- Autoclave
Procedure
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文章信息
版权信息
© 2015 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Chun, C. L., Nelson, M. S. and Sadowksy, M. J. (2015). Quantitative Evaluation of Competitive Nodulation among Different Sinorhizobium Strains. Bio-protocol 5(15): e1555. DOI: 10.21769/BioProtoc.1555.
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分类
微生物学 > 微生物-宿主相互作用 > 细菌
微生物学 > 微生物细胞生物学 > 细胞分离和培养
植物科学 > 植物新陈代谢 > 氮
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