1: Dev Biol. 2000 Aug 1;224(1):20-8. 

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

Oates AC, Bruce AE, Ho RK.

Department of Molecular Biology, Princeton University, Princeton, New Jersey
08544, USA.

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. Copyright 2000 Academic Press.

PMID: 10898958 [PubMed - indexed for MEDLINE]


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