Plant Assays for Quantifying Ralstonia solanacearum Virulence
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Author:
Date:
2018-09-20
[Abstract] Virulence assays are powerful tools to study microbial pathogenesis in vivo. Good assays track disease development and, coupled with targeted mutagenesis, can identify pathogen virulence factors. Disease development in plants is extremely sensitive to environmental factors such as temperature, atmospheric humidity, and soil water level, so it can be challenging to standardize conditions to achieve consistent results. Here, we present optimized and validated experimental conditions and analysis methods for nine assays that measure specific aspects of virulence in the phytopathogenic bacterium Ralstonia solanacearum, using tomato as the model host plant.
[摘要] 毒力测定是研究体内微生物发病机制的有力工具。 良好的分析跟踪疾病发展,并结合定向诱变,可以识别病原体毒力因子。 植物的疾病发展对环境因素如温度,大气湿度和土壤水位极其敏感,因此标准化条件以获得一致的结果可能具有挑战性。 在这里,我们提出优化和验证的实验条件和分析方法的九个测定,测量植物病原细菌 Ralstonia solanacearum 的毒力的特定方面,使用番茄作为模型宿主植物。
【背景】 Ralstonia solanacearum 是一种土壤传播的细菌,在广泛的植物中引起细菌枯萎,并继续感染全球的新宿主(Hayward,1991; Elphinstone,2005; Wicker et al。 ,2007; Genin,2010; Weibel et al。,2016)。结果, R. solanacearum 是研究最深入的植物致病菌之一(Mansfield et al。,2012)。
R上。 solanacearum 可以长期存在于土壤或水库中(Alvarez et ...
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Quantification of Triphenyl-2H-tetrazoliumchloride Reduction Activity in Bacterial Cells
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Author:
Date:
2017-01-20
[Abstract] This protocol describes the use of the 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) salt to evaluate the cell redox potential of rhizobia cells. The production of brightly colored and insoluble 1,3,5-Triphenyltetrazolium formazan arising from TTC reduction is irreversible and can be easily quantified using a spectrophotometer. This protocol allows the production of reproducible results in a relatively short time for Sinorhizobium meliloti 1021 cells grown both in exponential and stationary phases. The results here presented show that the S. meliloti cells deriving from exponential-phase cultures had increased cell redox potential as compared to the ones deriving from stationary-phase cultures. This means that under exponential growth conditions the S. meliloti ...
[摘要] 该方案描述了使用2,3,5-三苯基-2H-四唑氯化物(TTC)盐来评估根瘤细胞的细胞氧化还原电位。由TTC还原产生的鲜色不溶性1,3,5-三苯基四唑甲The的生产是不可逆的,可以使用分光光度计轻松地定量。该方案允许在相对较短的时间内在指数阶段和固定阶段生长的中华根瘤菌中华根瘤菌1021细胞产生可重复的结果。这里提出的结果表明,与来自固定相培养物的细胞相比,从指数阶段培养物得到的猕猴桃细胞具有增加的细胞氧化还原电位。这意味着在指数增长条件下, meliloti 细胞产生更大量的TTC减少所需的还原等同物。
背景 TTC盐是一种水溶性和无色的化合物,可以还原成甲an,高度着色的化合物。甲醛的不可逆形成可以使用分光光度计进行定量。由于其性质和其降低的潜力,该四唑盐广泛用于真核生物和原核生物两者,作为细胞氧化还原活性,活力,药物敏感性和底物利用测定的指标(Byth等人, 2001; Hayashi等人,2003; Raut等人,2008; Lin等人,2008)。由于膜电位,四唑鎓盐的净正电荷有助于细胞摄取,从而允许其细胞内还原(Berridge等人,2005)。在原核生物中,TTC降低的主要研究涉及革兰氏阴性呼吸细菌大肠杆菌,而对Rhizobiacea 家族的成员只有少数研究报道。在这个协议中,这个家族最好的遗传特征成员之一, ...
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Seed Coat Permeability Test: Tetrazolium Penetration Assay
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Author:
Date:
2014-07-05
[Abstract] Seed coat permeability is important to study as it plays significant roles in seed dormancy, germination, and protection from pathogens. Here we describe a commonly used seed coat permeability test known as the tetrazolium penetration assay with a method to quantify the levels of permeability. Tetrazolium red is a cationic dye that is widely used in seed viability testing. Tetrazolium salts are amphipathic cations, which, after penetrating the dead cells of the seed coat, are reduced to red-colored insoluble precipitates made up of formazans by active dehydrogenases (NADH-dependent reductases) in the embryo of seeds (Berridge et al., 1996). The intensity of red coloration is directly proportional to the permeability of the seeds. The quantification involves extraction of ...
[摘要] 种子包衣渗透性对于研究是重要的,因为它在种子休眠,发芽和对病原体的保护中起重要作用。 在这里,我们描述了一种常用的种皮渗透性测试称为四唑渗透测定法与量化渗透性水平的方法。 四唑红是一种阳离子染料,广泛用于种子活力测试。 四唑盐是两亲性阳离子,其在穿透种皮的死细胞后,在种子胚中被活性脱氢酶(NADH依赖性还原酶)还原为由甲偶合物形成的红色不溶性沉淀物(Berridge等人 al。,1996)。 红色着色的强度与种子的渗透性成正比。 定量包括从孵育的种子中提取甲seeds,并在485nm下分光光度测定甲an提取物的吸光度。 注意:此协议已针对测试拟南芥种子进行了优化。
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