PUBLICATION

Too much interference: injection of double-stranded RNA has nonspecific effects in the zebrafish embryo

Authors

Oates, A.C., Bruce, A.E., and Ho, R.K.

ID

ZDB-PUB-000824-12

Date

2000

Source

Developmental Biology 224(1): 20-28 (Journal)

Registered Authors

Bruce, Ashley, Ho, Robert K., Oates, Andrew

Keywords

spadetail; tbx16; no tail; nieuwkoid; dsRNA; RNAi; zebrafish; embryogenesis

MeSH Terms

Zebrafish/embryology
Zebrafish/genetics*
In Situ Hybridization
Embryo, Nonmammalian
Phenotype
RNA, Double-Stranded/genetics*
RNA, Messenger/analysis
Animals
T-Box Domain Proteins/genetics
T-Box Domain Proteins/metabolism*
Microinjections
Gene Silencing
Zebrafish Proteins*

PubMed

10898958 Full text @ Dev. Biol.

Abstract

We have investigated the ability of double-stranded RNA (dsRNA) to inhibit gene expression in a vertebrate, the zebrafish, Danio rerio. Injection of dsRNA corresponding to the T-box gene tbx16/spadetail (spt) into early wild-type embryos caused a rapid and dramatic loss of tbx16/spt mRNA in the blastula. mRNAs from the papc, tbx6, and gata1 genes, which depend on tbx16/spt function for their expression, were reduced, apparently mimicking the spt mutant phenotype. However, mRNAs from a number of genes that are unaffected by the spt mutation, such as beta catenin, stat3, and no tail, were also lost, indicating that the "interference" effect of tbx16/spt dsRNA was not restricted to the endogenous tbx16/spt mRNA. We compared the effects of injecting dsRNA from the zebrafish tbx16/spadetail, nieuwkoid/bozozok, and Brachyury/no tail genes with dsRNA from the bacterial lacZ gene. In each case the embryos displayed a variable syndrome of abnormalities at 12 and 24 h postfertilization. In blind studies, we could not distinguish between the effects of the various dsRNAs. Consistent with a common effect of dsRNA, regardless of sequence, injection of dsRNA from the lacZ gene was likewise effective in strongly reducing tbx16/spt and beta catenin mRNA in the blastula. These findings indicate that, despite published reports, the current methodology of double-stranded RNA interference is not a practical technique for investigating zygotic gene function during early zebrafish development.

Genes / Markers

Figures