Quantification of Ethylene Production in Tomato Leaves Infected by Xanthomonas euvesicatoria
黄单胞菌感染番茄叶片后乙烯产生的定量测定
发布: 2016年02月05日第6卷第3期 DOI: 10.21769/BioProtoc.1723 浏览次数: 11059
评审: Zhaohui LiuMalou FraitureDaniel F. CaddellArsalan Daudi
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参见作者原研究论文
The authors used this protocol in:
Feb 2015
Abstract
Ethylene is a gaseous plant hormone controlling fruit ripening, flower opening, leaf senescence as well as abscission, and disease symptom development. Ethylene plays a critical role in the bacterial pathogen Xanthomonas euvesicatoria (X. euvesicatoria)-elicited symptom development in tomato. This protocol describes the measurement of ethylene gas produced by tomato leaves infected with X. euvesicatoria. Infected leaflets are placed in a glass tube for 30 min without sealing. The glass tubes are then capped with a septa stopper, and incubated for an hour. A 1 ml gas sample is removed from the tube using a syringe and then injected into a gas chromatograph to quantify ethylene gas levels. This protocol will be applicable for other plants with other pathogens with modifications.
Background
Materials and Reagents
- 1% Ethylene gas can (SCOTT Specialty Gases, catalog number: 01-04-855 )
- 1 L Tedlar® PLV Gas Sampling Bag w/Thermogreen® LB-2 Septa (Sigma-Aldrich, Supelco, catalog number: 24633 )
- 1.5 ml Microtubes (Corning, Axygen®, catalog number: MTC-150-C )
- 1 ml Syringes Without Needles (BD, catalog number: 309659 )
- 25 G Needle (BD, catalog number: 305122 )
- 15 ml Polypropylene Centrifuge Tubes (Greiner Bio-One GmbH, catalog number: 188271 )
- 16 mm diameter x 100 mm Glass tubes (VWR International, catalog number: 47729-576 )
- 18 mm diameter x 150 mm Glass tubes (VWR International, catalog number: 47729-583 )
- Suba-Seal® septa (Sigma-Aldrich, catalog number: Z124613 )
- Tomato plants (4-5 week-old) were grown in a greenhouse or growth chamber (16 h light, 25~28 °C)
- Xanthomonas euvesicatoria (Xcv) strain 85-10 wild type and mutants (for example, type III effector deletion mutants)
Note: Wild type strain is available upon request. - Magnesium chloride hexahydrate (Sigma-Aldrich, catalog number: M2393 )
- Peptone (BD, catalog number: 211677 )
- Yeast extract (BD, catalog number: 212750 )
- Agar (BD, catalog number: 214530 )
- Glycerol (Certified ACS) (Fisher Chemical, catalog number: G334 )
- Sodium hydroxide (Sigma-Aldrich, catalog number: 221465 )
- Distilled water
- NYGA medium (see Recipes)
- 10 mM MgCl2 (see Recipes)
Equipment
- 28 ºC Incubator (VWR International, catalog number: 414005-128 )
- Vortexer (Scientific Indrustries, model: Vortex-Genie 2 )
- Spectrophotometer (Amersham Biosciences, model: Ultrospec 3100 Pro )
- Gas chromatograph (GC) (Shimadzu Corporation, model: GC-8A )
Procedure
文章信息
版权信息
© 2016 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Kim, J., Stork, W. and Mudgett, M. B. (2016). Quantification of Ethylene Production in Tomato Leaves Infected by Xanthomonas euvesicatoria. Bio-protocol 6(3): e1723. DOI: 10.21769/BioProtoc.1723.
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分类
植物科学 > 植物免疫 > 病害生物测定
微生物学 > 微生物-宿主相互作用 > 细菌
生物化学 > 其它化合物 > 植物激素 > 吲哚
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