An LNA-based loss-of-function assay for micro-RNAs.
MicroRNAs (miRNAs) have recently emerged as being essential for development and for the control of cell proliferation/differentiation in various organisms. However, little is known about miRNA function and mode of action at the cellular level. We have designed a miRNA loss-of-function assay, based on chemically modified locked nucleic acids (LNA) antisense oligonucleotides and usable in tissue culture cells. We show that LNA/DNA mixed oligonucleotides form highly stable duplexes with miRNAs in vitro. Ex vivo, the target miRNA becomes undetectable in cells transfected with the antisense oligonucleotide. The effect is dose-dependent, long-lasting, and specific. Moreover, using a reporter assay, we show that antisense LNA/DNA oligonucleotides inhibit short non-coding RNAs at the functional level. Thus LNA/DNA mixmers represent powerful tools for functional analysis of miRNAs.
Naguibneva
I
I
CNRS FRE 2944, Ligue nationale contre le cancer, 7, rue Guy Moquet, 94801 Villejuif cedex, France. nirina@vjf.cnrs.fr
Ameyar-Zazoua
M
M
Nonne
N
N
Polesskaya
A
A
Ait-Si-Ali
S
S
Groisman
R
R
Souidi
M
M
Pritchard
L L
LL
Harel-Bellan
A
A
D002478:Cells, Cultured; D006801:Humans; D035683:MicroRNAs / Q000037:antagonists & inhibitors / Q000502:physiology*; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*
D035683:MicroRNAs; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; C477371:locked nucleic acid