Tol2-Mediated Delivery of miRNAs to the Chicken Otocyst Using Plasmid Electroporation.
The avian embryo has a well-documented history as a model system for the study of neurogenesis, morphogenesis, and cell fate specification. This includes studies of the chicken inner ear that employ in ovo electroporation, in conjunction with the Tol2 system, to yield robust long-term transgene expression. Capitalizing on the success of this delivery method, we describe a modified version of the Tol2 expression vector that readily accepts the insertion of a microRNA-encoding artificial intron. This offers a strategy to investigate the possible roles of different candidate microRNAs in ear development by overexpression. Here, we describe the general design of this modified vector and the electroporation procedure. This approach is expected to facilitate phenotypic screening of candidate miRNAs to explore their bioactivity in vivo.
Department of Neurobiology and Anatomy, University of Utah, Salt Lake City, UT, USA.
Michelle L
ML
Stoller
Department of Biological Sciences, Purdue University, 915 W. State Street, West Lafayette, IN, 47907-2054, USA. dfekete@purdue.edu.|Purdue Center for Cancer Research, Purdue University, West Lafayette, IN, USA. dfekete@purdue.edu.
Donna M
DM
Fekete
D000818:Animals; D002642:Chick Embryo; D007758:Ear, Inner / Q000166:cytology / Q000196:embryology*; D018274:Electroporation / Q000379:methods*; D018014:Gene Transfer Techniques; D035683:MicroRNAs / Q000235:genetics*; D009865:Oocytes / Q000254:growth & development; D010641:Phenotype; D010957:Plasmids / Q000008:administration & dosage* / Q000235:genetics; D019895:Transposases / Q000378:metabolism
D035683:MicroRNAs; D019895:Transposases
Auditory; Inner ear; Tol2 transposon; Vestibular; microRNA