Double-stranded RNA injection produces nonspecific defects in zebrafish

Dev Biol. 2001 Jan 1;229(1):215-23. doi: 10.1006/dbio.2000.9982.


We have investigated the ability of dsRNA to inhibit gene functions in zebrafish using sequences targeted to the maternal gene pouII-1, the transgene GFP, and an intron of the zebrafish gene terra. We found that embryos injected with all of these dsRNAs at approximately 7.5 pg/embryo or higher had general growth arrest during gastrulation and displayed various nonspecific defects at 24 h postfertilization, although embryonic development was unaffected before the midblastula stage. Reducing dsRNA concentration could alleviate the global defects. Injection of GFP dsRNA (7.5-30 pg/embryo) did not inhibit GFP expression in transgenic fish, although abnormal embryos were induced. Co-injection of GFP mRNA with either GFP or non-GFP dsRNA caused reduction of GFP expression. Whole-mount in situ hybridization clearly showed that embryos injected with dsRNA degraded co-injected and endogenous mRNA without sequence specificity, indicating that dsRNA has a nonspecific effect at the posttranscriptional level. It appears that RNAi is not a viable technique for studying gene function in zebrafish embryos.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Abnormalities, Multiple / chemically induced*
  • Animals
  • Animals, Genetically Modified
  • DNA-Binding Proteins / genetics
  • Dose-Response Relationship, Drug
  • Green Fluorescent Proteins
  • In Situ Hybridization
  • Luminescent Proteins / genetics
  • Microinjections
  • Octamer Transcription Factor-3
  • RNA Stability
  • RNA, Antisense / toxicity
  • RNA, Double-Stranded / toxicity*
  • RNA, Messenger / metabolism
  • RNA, Small Interfering
  • Transcription Factors / genetics
  • Transgenes
  • Zebrafish
  • Zebrafish Proteins*


  • DNA-Binding Proteins
  • Luminescent Proteins
  • Octamer Transcription Factor-3
  • Pou5f3 protein, zebrafish
  • RNA, Antisense
  • RNA, Double-Stranded
  • RNA, Messenger
  • RNA, Small Interfering
  • Transcription Factors
  • Zebrafish Proteins
  • dmrt2a protein, zebrafish
  • Green Fluorescent Proteins