常见问题


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微奥基因的EMSA技术专辑在我公司网站上放了许多年,相信给我们的客户带来不少帮助。我们现正在对有关内容进行系统更新。近几年,我们发现微奥基因的所有网络文字与图片资料在国内被一些公司肆意盗用。未经同意而收纳我公司说明书,产品介绍, 公司培训资料与我公司于客户的合同书的包括一些知名网站如百度百科,百度知道,豆丁书房等。我公司的产品说明被一些公司 一字不差的照搬。有的公司根本就直接冒名为微奥基因对外从事欺诈。对于更新后的EMSA专辑,我们不再在网上全部公开。新的专辑将编辑 成带有电子追踪符号的eBook形式,通过电子邮件形式免费发给有需要的客户。新的EMSA专辑包括以下内容;

A. What is EMSA, why need EMSA.
B. The techniques for detecting the activity of transcriptional factors.
(1) What is the transcriptional factors?

(2) Common techniques for detecting TF.

(3) Which method is for detecting TF?
(4) Types of EMSA used for detection.
C. EMSA Principle
D. EMSA, experimental basis
(1) What needed for radioactive EMSA?
(2) What needed for non-radioactive EMSA?
(3) 成功进行凝胶迁移实验,需要优化哪些因素?
(4) 常用的同源多核苷酸引物、序列及其来源?
(5) The consideration for using non-radioactive EMSA probes.
(6) What DNA-binding proteins can be detected from HeLa cells.
(7) How much DNA-binding protein and non-radioactive probe are needed for EMSA?

(8) Why need Poly(dI:dC)(dI:dC) in EMSA experiments?
(9) What are specific and non-specific Competitive(竞争) EMSA?

(10) Why need to perform non-specific and specific Competitive(竞争) EMSA?

 (10)
(10) Why need Super-Shift/Supershift (超迁移)EMSA?

What needed for performing Supershift (超迁移)EMSA?

(11) 用什么凝胶条件可以将TF/探针复合物和游离的探针分开?
(12) Is non-radioactive EMSA less sensitive than that of radioactive EMSA?
(13) Whe need to prerun the gel for EMSA experiment?
E. The consideration for choosing EMSA reagents.
(1) 比较产品的成熟程度
(2) 是否供应配套产品
(3) Do not use the reagents/components from deferent supplies.

(4) 了解公司的技术支持力量
(5) There is any customer using that product.
(6) Are the customer satisfied with the product they used? 
F. What kinds of EMSA products on the market?
G. EMSA kits and products from 微奥基因?
H. How to perform a successful EMSA?
(1) Basic theory/knowledge about EMSA are required.
(2) A skill for performing vertical electrophoresis.

(3) A well designed subject or experiment.
(4) 需要仔细查看文献资料
(5) Good EMSA reagents are required for a successful EMSA?
(6) Equipment/devices for gel electrophoresis, imaging, UV-crosslinker etc. are needed for EMSA.
(7) High quality of protein sample
(8) A good negative/positive controls can save time, locate DNA-RNA-binding protein bands, and troubleshooting the problems.

9) Follow the operation manual provided by manufacturers.
(10) knowledge to know how to explain the result from EMSA.
(11) An ability to solve any possible problems.
I. The consideration of probe design and preparation.
(1) concerning about probe sequence.

(2) the position for labeling.
(3) How to purify the probes labeled with non-radioactive molecules?
(4) How to check the efficiency of probe labeling?
L. How to prepare samples for EMSA.

1) Quality of samples required for EMSA.

(2) Why are nuclear extracts needed for EMSA.

(3) The consideration of EMSA sample preparation.
(4) Can the protein expressed in titu be used for EMSA.
(5) How to shipping samples for EMSA services.
M. How to analyze EMSA results.

(1) 为什么EMSA带谱看不像Western Blot清晰?
(2) EMSA测定为什么出现非特异性结合带? 
(3) 为什么EMSA的结果有时不好分析?
(4) 没有阴性与阳性性对照时,如何对TF进行鉴定?
(5) 为什么TF的位置不能够通过分子量的大小来判断?
(6) 为什么EMSA测定中看到与阳性对照的位置不同?
(7) 如何在一个特定的复合物中确定亚单位蛋白的种类?
N. 常见失败原因分析
(1) 制备的样品中没有活化的转录因子。
(2) 核酸结合蛋白在测定前被降解。
(3) 核酸结合蛋白的天然状况被改变。
(4) 实验设计缺陷。
(5) 网上的探针序列不好直接用于nrEMSA。
(6) 实验条件受限(查看需要的条件)。
(7) EMSA试剂问题。
O. 参考资料
 

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