The use of in ovo electroporation for the rapid analysis of neural-specific murine enhancers.
The identification and characterization of DNA sequences necessary for proper gene expression have provided insights into gene regulation and generated tools useful for further experimentation. Studies of developmentally regulated genes have demonstrated how transcription factors interact at enhancers to generate restricted patterns of expression during embryogenesis. In vertebrates, the pursuit of such studies has relied on the generation of transgenic mice and thus has been limited by the time and expense required generating and characterizing these mice. The recently developed technique of in ovo electroporation allows the rapid introduction of exogenous DNA into developing chicken embryos. Here we have used this technique to introduce DNA containing murine enhancer/reporter constructs into cells of the chicken neural tube, resulting in appropriate expression of the reporter. This technique has the potential to greatly reduce the effort involved in the study of vertebrate enhancers. Furthermore, we have characterized factors such as timing of electroporation, concentration of DNA, and choice of basal promoters and found that they can influence the degree to which expression of enhancer constructs reflects endogenous gene expression.
Timmer
J
J
Molecular Biology Program and Howard Hughes Medical Institute, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Johnson
J
J
Niswander
L
L
D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins; D002642:Chick Embryo; D018274:Electroporation / Q000379:methods*; D004742:Enhancer Elements, Genetic; D020869:Gene Expression Profiling; D051379:Mice; D009419:Nerve Tissue Proteins / Q000235:genetics; D009474:Neurons; D011401:Promoter Regions, Genetic; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics
D051792:Basic Helix-Loop-Helix Proteins; D009419:Nerve Tissue Proteins; D014157:Transcription Factors; C491183:NEUROG2 protein, human; C102938:Neurog2 protein, mouse; C486336:Neurog1 protein, mouse