TY  - JOUR
TI  - Isolation of teleost primordial germ cells using flow cytometry 
AU  - Goto-Kazeto, Rie
AU  - Saito, Taiju
AU  - Takagi, Misae
AU  - Arai, Katsutoshi
AU  - Yamaha, Etsuro
T2  - The International Journal of Developmental Biology
AB  - Primordial germ cells (PGCs) generate gametes, the only cells that can transmit genetic information to the next generation. A previous report demonstrated that a fusion construct of green fluorescent protein (gfp) and zebrafish nos 1 3’UTR mRNA could be used to label PGCs in a number of fish species. Here, we sought to exploit this labeling strategy to isolate teleost PGCs by flow cytometry (FCM), and to use these isolated PGCs to examine germ cell migration to the gonadal region. In zebrafish, medaka and goldfish, the PGCs were labeled by injecting the gfp-nos1 3’UTR mRNA into 1- 4 cell embryos. When the embryos had developed to the somitogenesis or later stages, they were enzymatically disaggregated and GFP positive cells isolated using FCM. PGCs in the different species clustered in the same segments of the FCM scatter diagrams for total embryonic cells produced by plotting the forward scatter intensity against GFP intensity. In situ hybridization showed that the sorted zebrafish cells expressed vasa RNA in their cytoplasm, suggesting that they were PGCs. When the migration ability of the sorted cells from zebrafish was examined in an in vivo transplantation experiment, approximately 30% moved to the gonadal region of host embryos. These observations demonstrate that PGCs can be isolated without use of transgenic fishes and that the isolated PGCs retain the ability to migrate. Our data indicate that this technique will be of value for isolating PGCs from a range of fish species. 
PY  - 2010
DO  - 10.1387/ijdb.092914rg
VL  - 54
IS  - 10
SP  - 1487
EP  - 1492
J2  - Int. J. Dev. Biol.
LA  - en
SN  - 0214-6282
SN  - 1696-3547
UR  - https://ijdb.ehu.eus/article/092914rg
Y2  - 2024/11/21/18:05:18
ER  -