Using FAM labeled DNA oligos to do RNA electrophoretic mobility shift assay

被引:0
作者
Kun Wang
Ya Gao
Xiaojue Peng
Guohua Yang
Feng Gao
Shaoqing Li
Yingguo Zhu
机构
[1] Wuhan University,Key Laboratory of MOE for Plant Developmental Biology, College of Life Sciences
来源
Molecular Biology Reports | 2010年 / 37卷
关键词
EMSA; RNA–protein interactions; Fluorescence; Fluorescence imaging systems;
D O I
暂无
中图分类号
学科分类号
摘要
The electrophoretic mobility shift assay is a useful tool to identify proteins and nucleic acids interactions. Traditionally, the nucleic acids fragments are end-labeled with 32P. We present here the use of fluorescent methodologies for studies of RNA in place of radioactivity. The method is highly sensitive and quantitative with the use of an infrared fluorescence imaging system. Fluorescently labeled primers can be used to monitor protein–RNA interactions by fluorescent mobility shift assays. The simplicity and validity of this approach may have more advantages than that of previous methods that traditionally used hazardous radioisotopes. This method was successfully tested in the study of the interactions between MS2 Coat Protein and its RNA target.
引用
收藏
页码:2871 / 2875
页数:4
相关论文
共 35 条
[1]  
Hellman LM(2007)Electrophoretic mobility shift assay (EMSA) for detecting protein–nucleic acid interactions Nat Protoc 2 1849-1861
[2]  
Fried MG(1993)Nonradioactive gel mobility shift assay using chemiluminescent detection Biotechniques 15 650-652
[3]  
Berger R(2004)A modified quantitative EMSA and its application in the study of RNA–protein interactions J Biochem Biophys Methods 60 85-96
[4]  
Duncan MR(2000)Use of biotin-labeled nucleic acids for protein purification and agarose-based chemiluminescent electromobility shift assays Anal Biochem 277 254-259
[5]  
Berman B(2006)Quantitative analysis of DNA–protein interactions using double-labeled native gel electrophoresis and fluorescence-based imaging Electrophoresis 27 3166-3170
[6]  
Li Y(2002)Design of a fluorescent electrophoretic mobility shift assay improved for the quantitative and multiple analysis of protein–DNA complexes Biosci Biotechnol Biochem 66 2732-2734
[7]  
Jiang Z(2000)A fluorescence based non-radioactive electrophoretic mobility shift assay J Biotechnol 78 163-170
[8]  
Chen H(2007)A simple fluorescent method for detecting mismatched DNAs using a MutS-fluorophore conjugate Biosens Bioelectron 22 1376-1381
[9]  
Ma W-J(1997)RNA recognition by the MS2 phage coat protein Semin Virol 8 176-185
[10]  
Rodgers JT(undefined)undefined undefined undefined undefined-undefined