1 mm electroporation cuvette

Company: Genesee Scientific

Catalog#: 40-100

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Ratiometric Measurement of Protein Abundance after Transient Expression of a Transgene in Nicotiana benthamiana

Author:

Aashima Khosla and David C. Nelson,

Date:

2020-09-05

[Abstract] Ratiometric reporters are tools to dynamically measure the relative abundance of a protein of interest. In these systems, a target protein fused to a fluorescent or bioluminescent reporter is expressed with fixed stoichiometry to a reference protein fused to a second reporter. Both fusion proteins are encoded on a single transcript but are separated during translation by a 2A “self-cleaving” peptide. This approach enables changes in the relative abundance of a target protein to be detected sensitively, reducing variability in expression of the ratiometric reporter transgene that may occur across different tissues or transformation events. We recently developed a set of Gateway-compatible plant transformation vectors termed pRATIO that combine a variety of promoters, fluorescent and ... [摘要] [摘要] 比率报告基因是动态测量目标蛋白质相对丰度的工具。在这些系统中，与荧光或生物发光报告基因融合的靶蛋白以固定的化学计量比与与第二报告基因融合的参考蛋白表达。两种融合蛋白均编码在单个转录本上，但在翻译过程中被2A“自切割”肽隔开。这种方法使目标蛋白质的相对丰度的变化能够被敏感地检测到，从而减少了可能在不同组织或转化事件中发生的比例报告基因转基因表达的变异性。我们最近开发了一套与网关兼容的植物转化载体，称为pRATIO 结合了多种启动子，荧光和生物发光报告基因以及源自口蹄疫病毒的2A肽。在这里，我们详细描述了如何使用双荧光比率记者pRATIO3212检查靶蛋白的瞬时表达后的相对丰度烟草本塞姆氏叶。对于此示例，我们分析了响应于karrikins 和rac -GR24 处理的拟南芥MAX2 1（SMAX1）蛋白抑制子的降解。该方案提供了一种简单，快速，易于扩展的方法，用于体内分析农杆菌浸润的烟草叶片组织中的相对蛋白质丰度。

[背景] Karrikins （KARs ）是在烟雾中发现的丁烯内酯化合物，可刺激拟南芥种子萌发并增强幼苗的光形态发生（Flematti ，2004； Nelson 等，2009、2010 和2012）。拟南芥中的KAR响应需要α/β- 水解酶KARRIKIN对光的敏感性/低敏感性（KAI2 / HTL）（Sun and Ni，2011; ...

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1 mm electroporation cuvette

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