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Hokkaido University Collection of Scholarly and Academic Papers >
Graduate School of Fisheries Sciences / Faculty of Fisheries Sciences >
Peer-reviewed Journal Articles, etc >
Production of loach (Misgurnus anguillicaudatus) germ-line chimera using transplantation of primordial germ cells isolated from cryopreserved blastomeres
| Title: | Production of loach (Misgurnus anguillicaudatus) germ-line chimera using transplantation of primordial germ cells isolated from cryopreserved blastomeres |
| Authors: | Yasui, G.S. Browse this author | | Fujimoto, T. Browse this author | | Sakao, S. Browse this author | | Yamaha, E. Browse this author →KAKEN DB | | Arai, K. Browse this author →KAKEN DB |
| Keywords: | Aquaculture | | Cryobank | | Embryo | | Gamete | | Germ cell | | Germplasm | | Loach | | Teleost |
| Issue Date: | 1-Aug-2011 |
| Publisher: | American Society of Animal Science |
| Journal Title: | Journal of animal science |
| Volume: | 89 |
| Issue: | 8 |
| Start Page: | 2380 |
| End Page: | 2388 |
| Publisher DOI: | 10.2527/jas.2010-3633 |
| PMID: | 21398566 |
| Abstract: | An efficient procedure for cryopreservation of fish blastomeres followed by restoration through germ-line chimera formation was established. Blastomeres of the loach (Misgurnus anguillicaudatus) were cryopreserved in 250-µL straws in Eagle’s minimum essential medium (MEM) with various concentrations of dimethyl-sulfoxide (DMSO; 0, 5, 10, 15, and 20%), and the best concentration was combined with glycerol (1, 2, and 4%) and external cryoprotectants (1 or 2% sucrose; 2, 5, or 10% fetal bovine serum; 1 or 2% bovine serum albumin). Post-thaw viability of the blastomeres was used to optimize cryopreservation conditions. Donor blastomeres were injected with zebrafish GFP-nos1 3’UTR mRNA and biotin dextran prior to cryopreservation in the optimal freeze medium. Host embryos were injected with zebrafish DsRed-nos1 3’UTR mRNA and reared to the blastula stage. Donor blastomeres were thawed at 25°C for 10 s and transplanted to the host embryos either immediately or after incubation for 16 h at 20°C. Donor and host primordial germ cell migration was visualized with fluorescent imaging during the early stages of embryogenesis, and also by histology in 4-d-old embryos. Transplantation of blastomeres immediately after thawing gave lower hatching rates (~3%) and generated a low percentage of germ-line chimeras (~1.1%). In contrast, incubation of cryopreserved sample for 16 h followed by transplantation of the GFP-positive blastomeres improved the hatching rate to 90%, and successfully produced presumable germ-line chimeras at a rate of 16.5%. The improved survival rates and germ-line chimerism may be an effective method for gene banking and subsequent reconstitution of endangered fish genotypes. |
| Type: | article (author version) |
| URI: | http://hdl.handle.net/2115/49756 |
| Appears in Collections: | 水産科学院・水産科学研究院 (Graduate School of Fisheries Sciences / Faculty of Fisheries Sciences) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)
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Submitter: 荒井 克俊
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