Using Quantitative Real-Time PCR to Detect MicroRNA Expression Profile During Embryonic Stem Cell Differentiation.
Methods Mol Biol
; 1622: 255-265, 2017.
Article
in En
| MEDLINE
| ID: mdl-28674814
ABSTRACT
Quantitative real-time PCR (qRT-PCR) is a reliable method to determine and monitor microRNA (miRNA) expression profiles in different cells, tissues, and organisms. Although there are several different strategies in performing qRT-PCR to determine miRNA expression, all of them have two steps in common reverse transcription for obtaining cDNA from mature miRNA sequencing and standard real-time PCR for amplification of cDNA. This chapter demonstrates the application of quantitative real-time PCR for determining miRNA expression profiles during mouse embryonic stem cell differentiation. In this method, a mature miRNA sequence is first reverse transcribed into a long cDNA with a 40-50 nt miRNA-specific stem-loop primer; then, a standard real-time PCR reaction is performed for determining miRNA expression using a forward miRNA-specific primer and a universal reverse primer.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Cell Differentiation
/
Gene Expression Regulation, Developmental
/
MicroRNAs
/
Embryonic Stem Cells
/
Transcriptome
Limits:
Animals
Language:
En
Journal:
Methods Mol Biol
Journal subject:
BIOLOGIA MOLECULAR
Year:
2017
Document type:
Article
Affiliation country:
United States