Submitted December 12, 2007
Returned for revision January 9, 2008
Accepted April 25, 2008
Reproductive Technology
Increasing the Cell Number of Host Tetraploid Embryos Can Improve the Production of Mice Derived from Embryonic Stem Cells
Hiroshi Ohta *,
Yuko Sakaide ,
Kazuo Yamagata ,
and
Teruhiko Wakayama
* To whom correspondence should be addressed. E-mail: ohta{at}cdb.riken.go.jp.
Abstract
Tetraploid (4n) embryo complementation assay has shown that embryonic stem (ES) cells alone are capable of supporting embryonic development (ES mouse), allowing the recovery of mouse lines directly from cultured ES cell lines. Although the advantages of this technique are well recognized, it is still inefficient for generating ES mice. In this study, we investigated the effects of cell number of host 4n embryos on the production of ES mice. Four independent ES cell lines (2 general ES cell lines and 2 nuclear transfer-derived ES cell lines [ntES]) were used and each cell line was aggregated with single (1x) to triple (3x) host 4n embryos. We found that birthrate of ES mice using 1x 4n embryos was quite low (0-2%) regardless of cell line, whereas approximately 6-14% of embryos developed to full-term in the case of 3x 4n embryos, except for one cell line. Contamination of host 4n cells in ES mice was quite rare even if they were delivered from 3x 4n host embryos, comparable to that generated using general methods. These results demonstrate that the use of 3x 4n embryos is an effective procedure for generating ES mice. Our technique described here will be applicable to any ES cell lines including general ES cell lines used for gene targeting.
Key words:
Embryo •
Developmental biology •
embryonic stem cells •
tetraploid complementation assay •
trophectoderm
