TY - JOUR
T1 - Tet-on inducible system combined with in ovo electroporation dissects multiple roles of genes in somitogenesis of chicken embryos
AU - Watanabe, Tadayoshi
AU - Saito, Daisuke
AU - Tanabe, Koji
AU - Suetsugu, Rinako
AU - Nakaya, Yukiko
AU - Nakagawa, Shinichi
AU - Takahashi, Yoshiko
N1 - Funding Information:
We thank Dr. S. F. Gilbert for careful reading of the manuscript. We are also grateful to Dr. W. Hillen, Dr. A. Buchburger Dr. K. Kaibuchi and Dr. H. Nakamura for the kind gifts of rtTA2 S -M2, cMeso-1, RhoA and Pax2, respectively. This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan.
PY - 2007/5/15
Y1 - 2007/5/15
N2 - The in ovo electroporation technique in chicken embryos has enabled investigators to uncover the functions of numerous developmental genes. In this technique, the ubiquitous promoter, CAGGS (CMV base), has often been used for overexpression experiments. However, if a given gene plays a role in multiple steps of development and if overexpression of this gene causes fatal consequences at the time of electroporation, its roles in later steps of development would be overlooked. Thus, a technique with which expression of an electroporated DNA can be controlled in a stage-specific manner needs to be formulated. Here we show for the first time that the tetracycline-controlled expression method, "tet-on" and "tet-off", works efficiently to regulate gene expression in electroporated chicken embryos. We demonstrate that the onset or termination of expression of an electroporated DNA can be precisely controlled by timing the administration of tetracycline into an egg. Furthermore, with this technique we have revealed previously unknown roles of RhoA, cMeso-1 and Pax2 in early somitogenesis. In particular, cMeso-1 appears to be involved in cell condensation of a newly forming somite by regulating Pax2 and NCAM expression. Thus, the novel molecular technique in chickens proposed in this study provides a useful tool to investigate stage-specific roles of developmental genes.
AB - The in ovo electroporation technique in chicken embryos has enabled investigators to uncover the functions of numerous developmental genes. In this technique, the ubiquitous promoter, CAGGS (CMV base), has often been used for overexpression experiments. However, if a given gene plays a role in multiple steps of development and if overexpression of this gene causes fatal consequences at the time of electroporation, its roles in later steps of development would be overlooked. Thus, a technique with which expression of an electroporated DNA can be controlled in a stage-specific manner needs to be formulated. Here we show for the first time that the tetracycline-controlled expression method, "tet-on" and "tet-off", works efficiently to regulate gene expression in electroporated chicken embryos. We demonstrate that the onset or termination of expression of an electroporated DNA can be precisely controlled by timing the administration of tetracycline into an egg. Furthermore, with this technique we have revealed previously unknown roles of RhoA, cMeso-1 and Pax2 in early somitogenesis. In particular, cMeso-1 appears to be involved in cell condensation of a newly forming somite by regulating Pax2 and NCAM expression. Thus, the novel molecular technique in chickens proposed in this study provides a useful tool to investigate stage-specific roles of developmental genes.
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U2 - 10.1016/j.ydbio.2007.01.042
DO - 10.1016/j.ydbio.2007.01.042
M3 - Article
C2 - 17359965
AN - SCOPUS:34247552289
VL - 305
SP - 625
EP - 636
JO - Developmental Biology
JF - Developmental Biology
SN - 0012-1606
IS - 2
ER -