title: In vivo testing of microRNA-mediated gene knockdown in zebrafish.
abstract: The zebrafish (Danio rerio) has become an attractive model for human disease modeling as there are a large number of orthologous genes that encode similar proteins to those found in humans. The number of tools available to manipulate the zebrafish genome is limited and many currently used techniques are only effective during early development (such as morpholino-based antisense technology) or it is phenotypically driven and does not offer targeted gene knockdown (such as chemical mutagenesis). The use of RNA interference has been met with controversy as off-target effects can make interpreting phenotypic outcomes difficult; however, this has been resolved by creating zebrafish lines that contain stably integrated miRNA constructs that target the desired gene of interest. In this study, we show that a commercially available miRNA vector system with a mouse-derived miRNA backbone is functional in zebrafish and is effective in causing eGFP knockdown in a transient in vivo eGFP sensor assay system. We chose to apply this system to the knockdown of transcripts that are implicated in the human cardiac disorder, Long QT syndrome.
authors.affiliation: School of Biological Sciences, University of Auckland, Auckland, New Zealand.
authors.forename: Ivone Un San
authors.initials: IU
authors.lastname: Leong
authors.forename: Chuan-Ching
authors.initials: CC
authors.lastname: Lan
authors.forename: Jonathan R
authors.initials: JR
authors.lastname: Skinner
authors.forename: Andrew N
authors.initials: AN
authors.lastname: Shelling
authors.forename: Donald R
authors.initials: DR
authors.lastname: Love
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D004195:Disease Models, Animal; D004625:Embryo, Nonmammalian; D051638:Ether-A-Go-Go Potassium Channels / Q000235:genetics / Q000378:metabolism; D055785:Gene Knockdown Techniques / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008133:Long QT Syndrome; D051379:Mice; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics*; D034622:RNA Interference; D015027:Zebrafish; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: C527141:Erg protein, zebrafish; D051638:Ether-A-Go-Go Potassium Channels; D035683:MicroRNAs; D029961:Zebrafish Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins