Abstract
We describe the derivation and characterization of three novel human embryonic stem (hES) cell lines (YT1, YT2, YT3). One hES line (YT1) was obtained from six discarded blastocysts in a culture medium supplemented with 12 ng/ml basic fibroblast growth factor (bFGF), and two lines (YT2, YT3) were obtained from three discarded blastocysts in the same medium but supplemented with 16 ng/ml bFGF. These cell lines were derived by partial or whole embryo culture followed by further expansion after manual dissection of the passaged cells. These cells were passaged continuously for more than 6 or 8 months and possessed all of the typical features of pluripotent hES cell lines, such as typical morphological characteristics and the expression of hES-specific markers (TRA-1-60, TRA-1-81, SSEA-4, SSEA-3, alkaline phosphatase, Oct4, Nanog) and pluripotency-related genes (Oct4, Nanog, TDGF1, Sox2, EBAF, Thy-1, FGF4, Rex1). The lines maintained normal karyotypes after long-term cultivation. The karyotype of YT1 and YT3 was 46, XX, and that of YT2 was 46, XY. Pluripotency was confirmed by in vitro and in vivo differentiation, and genetic identity was demonstrated by DNA fingerprinting. Our results indicate that higher concentrations of bFGF at the early culture stage support efficient the hES cell derivation.
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Acknowledgments
We thank Prof. Yao Weidong for technical assistance in the histological analysis of teratoma sections, Hou Nanying for karyotype analysis, and Dr. TG Cooper for language editing. This research was supported by grants from the National Basic Research Program of China (973 Program, 2005CB522705-2), National High Technology Research and Development Program of China (863 program, 2003AA205008), Shandong Province Key Project of China (021100104), and Yantai Science and Technology Project (2009107).
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Y. Wang and C. Xu contributed equally to this work.
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Wang, Y., Xu, C., Wang, H. et al. Efficient derivation of human embryonic stem cell lines from discarded embryos through increases in the concentration of basic fibroblast growth factor. Human Cell 25, 16–23 (2012). https://doi.org/10.1007/s13577-011-0039-7
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DOI: https://doi.org/10.1007/s13577-011-0039-7