High-throughput real-time quantitative reverse transcription PCR.
Curr Protoc Mol Biol
; Chapter 15: Unit 15.8, 2006 Feb.
Article
em En
| MEDLINE
| ID: mdl-18265376
ABSTRACT
Extensive detail on the application of the real-time quantitative polymerase chain reaction (QPCR) for the analysis of gene expression is provided in this unit. The protocols are designed for high-throughput, 384-well-format instruments, such as the Applied Biosystems 7900HT, but may be modified to suit any real-time PCR instrument. QPCR primer and probe design and validation are discussed, and three relative quantitation methods are described the standard curve method, the efficiency-corrected DeltaCt method, and the comparative cycle time, or DeltaDeltaCt method. In addition, a method is provided for absolute quantification of RNA in unknown samples. RNA standards are subjected to RT-PCR in the same manner as the experimental samples, thus accounting for the reaction efficiencies of both procedures. This protocol describes the production and quantitation of synthetic RNA molecules for real-time and non-real-time RT-PCR applications.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
RNA
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Reação em Cadeia da Polimerase Via Transcriptase Reversa
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Pesquisa Biomédica
Idioma:
En
Revista:
Curr Protoc Mol Biol
Ano de publicação:
2006
Tipo de documento:
Article
País de afiliação:
Estados Unidos