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| Isolation, Culture and Characterization of Chicken Primordial Germ Cells |
| TANG Xin-yan;ZENG Wei-dong;MI Yu-ling;LIU Hong-yun;ZHANG Cai-qiao |
| College of Animal Sciences, Zhejiang University, Hangzhou 310029, China |
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Abstract Primordial germ cells (PGCs) were isolated from the genital ridges of Avian chicken (Gallus domesticus) embryos at 19th stage and purified by Ficoll density-gradient centrifugation. PGCs were co-cultured with somatic cells in primary culture and subculture. Identification of PGCs was carried out by histochemical methods including alkaline phosphatase (AKP) and periodic acid-Schiff (PAS) staining, and proliferating PGCs in subculture was demonstrated by immunocytochemistry of proliferating cell nuclear antigen (PCNA). Meanwhile, proliferating activity of PGCs was compared under different culture conditions among 5% ~20% fetal calf serum (FCS), ITS (M199+10 μg/mL insulin+5 μg/mL transferrin + 3×10-8 mol/L selenite, conditioned medium (CM), 15%FCS+ITS, 15%FCS+40% CM. The results showed that the cultured PGCs were positive for AKP and PAS staining and displayed intensive proliferating activity by PCNA. PGCs grew better without centrifugation than those with centrifugation. PGCs formed larger colonies in media with 5%FCS or ITS than those in other media. The above results indicate that 5%FCS or ITS-supplemented media can be a culture system for PGC proliferation in the somatic cell- PGC coculture, besides the embryonic fibroblast feed layer.
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Received: 04 April 2005
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Corresponding Authors:
ZHANG Cai-qiao
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