Fisheries and Aquatic Sciences

The Korean Society of Fisheries and Aquatic Science (한국수산과학회)
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Simultaneous and Systemic Knock-down of Big Defensin 1 and 2 gene Expression in the Pacific Oyster Crassostrea gigas using Long Double-stranded RNA-mediated RNA Interference

  • Jee, Bo Young (Aquatic Life Disease Control Division, National Fisheries Research and Development Institute) ;
  • Kim, Min Sun (Department of Aquatic Life Medicine, Pukyong National University) ;
  • Cho, Mi Young (Aquatic Life Disease Control Division, National Fisheries Research and Development Institute) ;
  • Lee, Soon Jeong (Aquatic Life Disease Control Division, National Fisheries Research and Development Institute) ;
  • Park, Myung Ae (Aquatic Life Disease Control Division, National Fisheries Research and Development Institute) ;
  • Kim, Jin Woo (Pathology division, National Fisheries Research and Development Institute) ;
  • Choi, Seung Hyuk (Department of Aquatic Life Medicine, Pukyong National University) ;
  • Jeong, Hyun Do (Department of Aquatic Life Medicine, Pukyong National University) ;
  • Kim, Ki Hong (Department of Aquatic Life Medicine, Pukyong National University)
  • Received : 2013.06.28
  • Accepted : 2014.04.16
  • Published : 2014.09.30
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Abstract

RNA interference (RNAi)-mediated transcriptional knock-down of Crassostrea gigas big defensin 1 and 2 genes (Cg-BigDef1 and Cg-BigDef2) was investigated. The cDNA sequences of Cg-BigDef1 and Cg-BigDef2 were identical, excluding an additional fragment of 20 nucleotides in Cg-BigDef1; thus, a long double-stranded RNA (dsRNA) targeting the mRNA of Cg-BigDef2 effectively downregulated both Cg-BigDef2 and Cg-BigDef1. In addition, long dsRNA targeting green fluorescent protein (GFP) did not affect transcription of the two big defensin genes. These results suggest that the transcriptional downregulation of Cg-BigDef1 and Cg-BigDef2 was mediated by sequence-specific RNA interference (RNAi). Despite injection of long dsRNA targeting Cg-BigDef2 into only the adductor muscle, knock-down of Cg-BigDef1 and Cg-BigDef2 was observed in the adductor muscle, hemocytes, mantle, and gills, suggestive of systemic spread of RNAi in C. gigas. Furthermore, the inhibitory effect of dsRNA persisted until 72 h post-injection, indicative of a long-lasting RNAi-mediated knock-down of target genes.

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