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Properties of gene knockdown system by vector‐based si RNA in zebrafish
Author(s) -
Shinya Minori,
Kobayashi Kayo,
Masuda Aki,
Tokumoto Mika,
Ozaki Yuichi,
Saito Kenji,
Kawasaki Toshihiro,
Sado Yukiko,
Sakai Noriyoshi
Publication year - 2013
Publication title -
development, growth and differentiation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.864
H-Index - 66
eISSN - 1440-169X
pISSN - 0012-1592
DOI - 10.1111/dgd.12091
Subject(s) - rna interference , gene knockdown , small hairpin rna , biology , gene silencing , small interfering rna , rna silencing , gene , zebrafish , microrna , microbiology and biotechnology , rna , trans acting sirna , genetics
RNA interference (RNAi) has emerged as a powerful tool to silence specific genes. Vector‐based RNAi systems have been developed to downregulate targeted genes in a spatially and temporally regulated fashion both in vitro and in vivo . The zebrafish ( D anio rerio ) is a model animal that has been examined based on a wide variety of biological techniques, including embryonic manipulations, forward and reverse genetics, and molecular biology. However, a heritable and tissue‐specific knockdown of gene expression has not yet been developed in zebrafish. We examined two types of vector, which produce small interfering RNA (siRNA), the direct effector in RNAi system; microRNA (miRNA) process mimicking vectors with a promoter for RNA polymerase II and short hairpin RNA (shRNA) expressing vector through a promoter for RNA polymerase III. Though gene‐silencing phenotypes were not observed in the miRNA process mimicking vectors, the transgenic embryos of the second vector (Tg( zU 6‐shGFP)), shRNA expressing vector for enhanced green fluorescence protein , revealed knockdown of the targeted gene. Interestingly, only the embryos from Tg( zU 6‐shGFP) female but not from the male fish showed the downregulation. Comparison of the quantity of siRNA produced by each vector indicates that the vectors tested here induced siRNA, but at low levels barely sufficient to silence the targeted gene.