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左眼

金虫 (小有名气)

[求助] native page marker和还原剂问题 已有1人参与

1. 想知道不同条件下蛋白聚集情况。计算等电点在5.6,应该选择什么样的marker? 因为蛋白在SDS-PAGE上显示的是单体的分子量,可以用还原加热变性的蛋白做marker吗?
2. 蛋白本身易聚集,所以在纯化过程中一直有还原剂。这样可以跑native胶吗?native page能耐受的还原剂是什么剂量?
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zhouxfd

木虫 (正式写手)

【答案】应助回帖

感谢参与,应助指数 +1
1escription

The NativeMark™ Unstained Protein Standard is a ready-to-use protein standard designed for molecular weight estimation of proteins using native gel electrophoresis. Features of this marker:

• Versatile—recommended for use with Novex® NativePAGE™ Bis-Tris, gradient Tris-Glycine, or NuPAGE® Novex® Tris-Acetate gels
• Comprehensive—contains markers with a wide range of high molecular weights, providing 8 protein bands in the range of 20–1,200 kDa
• Convenient—no need to add sample buffer, just load and run
• Compatible—can be visualized using Coomassie®, silver, or fluorescent stains after electrophoresis, or with Ponceau S, Coomassie®, or other membrane stains after Western transfer

Electrophoresis under native (nondenaturing) conditions maintains the native protein conformation, cohesion of subunits, and biological activity. During native electrophoresis, proteins are separated based on their charge to mass ratios.

2: 10-20mM DTT 应该没问题
2楼2017-07-19 12:48:40
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vv一笑

新虫 (初入文坛)

引用回帖:
2楼: Originally posted by zhouxfd at 2017-07-19 12:48:40
1escription

The NativeMark™ Unstained Protein Standard is a ready-to-use protein standard designed for molecular weight estimation of proteins using native gel electrophoresis. Features of ...

我想要分子量小一点的Marker,也是用于Native-PAGE实验,大概10-100kDa的,知道哪里能订购吗
3楼2018-06-29 15:19:28
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