Primer Extension Analysis Using AMV Reverse Transcriptase
使用AMV逆转录酶进行引物延伸分析
发布: 2012年07月20日第2卷第14期 DOI: 10.21769/BioProtoc.240 浏览次数: 10561
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参见作者原研究论文
The authors used this protocol in:
Dec 2011
Abstract
Primer extension analysis is a useful method to determine the transcription start point or a processing site on an RNA molecule. It can also allow a quantitative measurement of an RNA species.
Keywords: 5' end mapping (5’端映射)
Reverse Transcription (反转录)
Primer extension (引物延伸)
Background
Materials and Reagents
- AMV reverse transcriptase from the avian myeloblastosis virus (Finnzyme, catalog number: F570 ) (other reverse transcriptases can also be used, by adapting the reaction buffer)
- T4 polynucleotide kinase + 10x reaction buffer (Biolabs, catalog number: M0201 )
- RNasin Plus RNase inhibitor (Promega Corporation, catalog number: N2611 )
- Glycogen (Acros organics, catalog number: 422950010 )
- NaCl
- Tris-HCl (pH 7.5)
- EDTA
- MgCl2
- EtOH
- dATP,dCTP,dGTP,dTTP (Promega Corporation, catalog number: U120D , U121D , U122D , U123D )
- γ-32P ATP (3,000 Ci/mmole, 10 μCi/μl) (PerkinElmer, catalog number: BLU502A )
- DTT (Promega Corporation, catalog number: P117B )
- 5x ss-Hybridization buffer (see Recipes)
- 1.25x RT buffer (see Recipes)
Equipment
- Incubator
Procedure
文章信息
版权信息
© 2012 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Putzer, H. (2012). Primer Extension Analysis Using AMV Reverse Transcriptase. Bio-protocol 2(14): e240. DOI: 10.21769/BioProtoc.240.
Download Citation in RIS Format
分类
分子生物学 > RNA > 转录
分子生物学 > RNA > qRT-PCR
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