LightShift Chemiluminescent EMSA Kit 化学发光试剂盒,LightShift Chemiluminescent EMSA Kit/化学发光EMSA试剂盒-试剂- bio-rad伯乐一级代理 |伯乐Aminex |伯乐电泳槽1658001| 伯乐ddPCR bio-rad授权代理商、biorad伯乐qpcr仪

LightShift Chemiluminescent EMSA Kit 化学发光试剂盒,LightShift Chemiluminescent EMSA Kit/化学发光EMSA试剂盒

2024-10-23

LightShift Chemiluminescent EMSA Kit/化学发光EMSA试剂盒

货号：20148

规格：100 reactions

价格：6320

产品类型：蛋白凝胶电泳

品牌：Thermo Fisher

Thermo Scientific LightShift化学发光EMSA试剂盒是一种非常强劲且灵敏的检测系统，通过进行电泳迁移率实验(EMSA)检测蛋白质-DNA结合相互作用。这个试剂盒包括用于建立和定制DNA结合反应的试剂，用于验证体系的一组对照DNA和蛋白提取物，作为探针去识别生物素标记靶DNA的稳定链霉亲和素-HRP偶联物，以及非常灵敏的用于检测的化学发光底物。LightShift EMSA检测的原理与蛋白免疫印迹类似。生物素末端标记的双链DNA与细胞核提取物或纯化的因子共同孵育，之后进行非变性凝胶电泳。然后将DNA快速（30分钟）转印至带正电的尼龙膜上，进行紫外交联，然后使用HRP标记的链霉亲和素-HRP偶联物检测，再与底物孵育。从标记到获得结果的整个实验流程可以在一天内完成。蛋白与DNA之间的相互作用对于DNA复制、重组和修复、转录和病毒组装等许多细胞进程具有关键调控作用。电泳迁移率实验(EMSA)是研究基因调控和确定蛋白:DNA之间相互作用的核心技术之一。EMSA技术基于一个实验发现：即在非变性聚丙烯酰胺凝胶或琼脂糖凝胶电泳过程中，蛋白:DNA复合物的迁移速度比自由的DNA分子更慢。由于蛋白结合使DNA的迁移速率发生了偏移或延迟，这个方法也被称为凝胶迁移或凝胶阻滞分析。在EMSA技术出现之前，蛋白:DNA之间相互作用主要是通过硝化纤维素膜过滤实验分析。整个实验中需要实验者准备的是已纯化的末端生物素标记的目标DNA，待检测的蛋白提取物，尼龙膜和基本电泳设备。目标DNA可在合成的同时进行5'或3'端生物素标记，或在合成之后使用Thermo Scientific生物素3'末端DNA标记试剂盒（产品货号89818）进行标记。用户可通过不同方法获得胞核、胞质或全细胞蛋白提取物，包括Thermo Scientific NE-PER 核蛋白和胞质蛋白提取试剂盒（产品货号78833）。

特点：

• 检测胞核抽提物中低丰度蛋白的理想选择• 灵敏度超越放射性和地高辛检测法• 兼容之前建立的常规DNA-蛋白互作结合条件• 包括EBNA对照系统，帮助新用户建立一个适当的检测体系，了解用于确定相互作用特异性的方法

数据：

图1. Procedure summary for the LightShift EMSA Kit

图2. More sensitive EMSA detection than other gel shift assay methodsComparison of the LightShift EMSA Kit to a popular digoxigenin-based EMSA kit and a radioactive method. Serial dilutions of a labeled DNA duplex were electrophoresed on a 6% polyacrylamide gel and detected according to the manufacturer’s instructions. Comparable sensitivity (<50 attomoles) was achieved with the LightShift EMSA Kit and radioactivity (2,000 cpm 32-P/fmol), although a significantly longer exposure was required for the 32P-labeled DNA. Equivalent exposures using the two chemiluminescent kits showed that the sensitivity of the LightShift EMSA Kit was approximately eight-fold greater than that of the digoxigenin kit.

图3. Better EMSA results in less timeComparison of the LightShift EMSA Kit to a popular digoxigenin-based EMSA kit and a radioactive method. A 22-bp duplex containing the binding sequence for the transcription factor Oct-1 was labeled for use in either the LightShift EMSA Kit, a digoxigenin-based EMSA kit or with 32P using T4 polynucleotide kinase (40,000 cpm/reaction) for use in traditional radioactive EMSA. Binding reactions were equivalent in that 20 fmol duplex was incubated with 6.8µg HeLa cell NE-PER Nuclear Extract (where indicated). The chemiluminescent kits were used according to the manufacturer’s instructions. For the radioactive EMSA, the gel was exposed directly to X-ray film using screens.

技术参数

组成成分

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LightShift Chemiluminescent EMSA Kit 化学发光试剂盒,LightShift Chemiluminescent EMSA Kit/化学发光EMSA试剂盒

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