Spatially and temporally controlled electroporation of early chick embryos.
The introduction of in ovo electroporation a decade ago has helped the chick embryo to become a powerful system to study gene regulation and function during development. Although this is a simple procedure for embryos of 2-d incubation, earlier stages (from laying to early neurulation, 0-1 d) present special challenges. Here we describe a robust and reproducible protocol for electroporation of expression vectors and morpholino oligonucleotides into the epiblast of embryos from soon after laying (stage XI) to stages 6-7 (early neurulation), with precise spatial and temporal control. Within 3 h, about 12 embryos can be electroporated and set up for culture by the New technique; the effects of morpholinos can be assessed immediately after electroporation, and robust overexpression from plasmid DNA is seen 2-3 h after electroporation. These techniques can be used for time-lapse imaging, gain- and loss-of-function experiments and studying gene regulatory elements in living embryos.
Voiculescu
Octavian
O
Department of Anatomy and Developmental Biology, University College London, Gower Street, London WC1E 6BT, UK. o.voiculescu@ucl.ac.uk
Papanayotou
Costis
C
Stern
Claudio D
CD
D000818:Animals; D002642:Chick Embryo; D018274:Electroporation / Q000379:methods*; D046149:Embryo Culture Techniques; D020869:Gene Expression Profiling / Q000379:methods*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D016376:Oligonucleotides, Antisense