ReadiLink Cy5 缺口平移 dsDNA 标记试剂盒

 

样品实验方案

简要概述

1. 准备 DNA 样本
2. 向试管中加入试剂
3. 短暂混合并离心
4. 在 15°C 下孵育 60 分钟
5. 将反应置于冰上，然后加入停止溶液并在 65°C 下加热
6. 使用前在冰上放置 5 分钟或在 4°C 下储存
7. 纯化标记的 DNA

 注：在开始实验之前，解冻所有成分。在开始标记过程之前，将所有试剂短暂涡旋至底部。

 

实验步骤

表 1. 各反应每管试剂组成

成分	规格
DNA样本	1 µg DNA 在无核酸酶水中稀释至终体积 34 µL
Nick Translation 缓冲液	5 µL
dNTP 混合物	5 µL
dTTP	2 µL
Cy5-dUTP 工作溶液	2 µL
DNA聚合酶I	1 µL
DNA酶I	1 µL
总容积	50 µL

可以优化Cy5-dUTP（组分 A）：dTTP（组分 E）的比例以获得佳标记条件。
可以优化孵育时间以获得更好的标记。 更长的孵育时间将有助于更多的标记，但可能会缩短终产品的尺寸。
1.向干净的（无核酸酶）0.5 mL 微型离心管或 0.2 mL PCR 管中，按表 1 中所示的顺序添加试剂。
2.通过短暂的涡旋和短暂的离心小心混合试剂。
3.将反应在 15°C 下孵育 60 分钟。
4.孵育后，将反应置于冰上。
5.要终止反应，请添加 5 µL 停止溶液并将样品加热至 65 °C。
6.使用前在冰上放置 5 分钟或在 4°C 下储存。
7.纯化标记的 DNA。

 

 

参考文献

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Tracing Baculovirus AcMNPV Infection Using a Real-Time Method Based on ANCHORTM DNA Labeling Technology.
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