ZFIN ID: ZDB-PUB-080722-6
Application of RNAi Technology to the Inhibition of Zebrafish GtHalpha, FSHbeta, and LHbeta Expression and to Functional Analyses
Huang, W.T., Hsieh, J.C., Chiou, M.J., Chen, J.Y., Wu, J.L., and Kuo, C.M.
Date: 2008
Source: Zoological science   25(6): 614-621 (Journal)
Registered Authors: Chen, Jyh-Yih, Huang, Wei-Tung, Wu, Jen-Leih
Keywords: zebrafish, RNA interference, knockdown, gonadotropin alpha, GtHα, common α, follicle-stimulating hormone beta, FSHβ, luteinizing hormone beta, LHβ
MeSH Terms:
  • Animals
  • Blotting, Southern
  • DNA Primers/genetics
  • Embryonic Development/drug effects*
  • Embryonic Development/genetics
  • Genetic Vectors/genetics
  • Gonadotropins, Pituitary/metabolism*
  • Gonadotropins, Pituitary/pharmacology
  • Green Fluorescent Proteins/metabolism
  • Microinjections
  • RNA/metabolism
  • RNA Interference*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Zebrafish/embryology*
  • Zebrafish/metabolism*
PubMed: 18624572 Full text @ Zool. Sci.
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ABSTRACT
Zebrafish (Danio rerio) were used as a model fish, and the technique of RNA interference (RNAi) was employed to knockdown three subunits of the gonadotropin alpha (GtHalpha, common alpha), follicle-stimulating hormone beta (FSHbeta), and luteinizing hormone beta (LHbeta) genes. Three short-hairpin RNA (shRNA) expression vectors and three mismatched shRNA expression vectors as controls for each subunit gene were constructed, and the depression efficiency was tested in vivo by microinjection; the RNA or protein expression levels of the GtH genes were monitored by RT-PCR, Southern blotting, and green fluorescent protein (GFP) analyses. Expression of GtH mRNA was obviously and more efficiently depressed by GtHalpha RNAi expression compared with the other two subunits. A GtHalpha morpholino analysis showed that the GtHalpha morpholino led to suppression of embryonic development and the production of embryonic mutants as a result of an injection of GtHalpha -shRNA. Taken together, these results show that GtHalpha-shRNA, which more efficiently targets RNAi, may have an essential role in the further development of sterility technology of transgenic fish for biosafety purposes.
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