标签：IRDye 700 EMSA 寡核苷酸结合使用

Mobility shift assay protocols can be easily converted to infrared fluorescent assays by replacing the existing DNA oligonucleotides with IRDye^® infrared dye end-labeled oligonucleotides. Binding and electrophoresis conditions are the same as any other EMSA detection method.

The Odyssey EMSA Kit, coupled with IRDye 700 EMSA oligonucleotides, is an excellent alternative method to radioisotopic and chemiluminescent detection methods for EMSA analysis and visualization^{1, 2}. Using IRDye EMSA reagents, assays can be completed in less than two hours with no gel transfer or film exposure. The gel doesn’t even need to be removed from the glass plates for imaging.

通过用 IRDye® 红外染料末端标记的寡核苷酸替换现有的 DNA 寡核苷酸，迁移率变化分析方案可以轻松转换为红外荧光分析。 结合和电泳条件与任何其他 EMSA 检测方法相同。

Odyssey EMSA 试剂盒与 IRDye 700 EMSA 寡核苷酸结合使用，是用于 EMSA 分析和可视化的放射性同位素和化学发光检测方法的绝佳替代方法1、2。使用 IRDye EMSA 试剂，可以在两小时内完成检测，无需凝胶转移或 胶片曝光。 凝胶甚至不需要从玻璃板上取出来进行成像。

Kit Components

• 10X Binding Buffer (100 mM Tris, 500 mM KCl, 10 mM DTT, pH 7.5), 500 μL
• 25 mM DTT, 2.5% Tween^® 20, 500 µL
• Poly (dI•dC), 1 µg/µL in 10 mM Tris, 1 mM EDTA, pH 7.5, 125 µL
• Sheared salmon sperm DNA, 0.5 µg/µL in 10 mM Tris, 1 mM EDTA, pH 7.5, 125 µL
• 50% Glycerol, 500 µL
• 1% NP-40, 500 µL
• 1 M KCl, 500 µL
• 100 mM MgCl₂, 500 µL
• 200 mM EDTA, pH 8.0, 500 µL
• 10X Orange Loading Dye, 500 µL
• 10X 结合缓冲液（100 mM Tris、500 mM KCl、10 mM DTT，pH 7.5），500 μL
  25 mM DTT, 2.5% Tween® 20, 500 µL
  Poly (dI•dC), 1 µg/µL 溶于 10 mM Tris, 1 mM EDTA, pH 7.5, 125 µL
  剪切的鲑鱼精子 DNA，0.5 µg/µL 溶于 10 mM Tris，1 mM EDTA，pH 7.5，125 µL
  50% 甘油，500 µL
  1% NP-40, 500 µL
  1 M 氯化钾，500 µL
  100 mM MgCl2，500 µL
  200 mM EDTA，pH 8.0，500 µL
  10X 橙色上样染料，500 µL

Example of an NIR Fluorescent EMSA

References

1. Li, Y., et al. (2005) Cancer Res. 65: 6934-6942. DOI: 10.1158/0008-5472.
2. Geddie, M.L., et al. (2005) J. Biol. Chem. 280(42):35641-6. DOI: 10.1074/jbc.M508149200