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Establishment of a transgenic quail model and an ex vivo culture system of yolk sac membrane endodermal epithelium cell for studying functions of individual genes in avian embryonic development
Author(s) -
Lin Han Jen,
Mersmann Harry,
Ding Shih Torng
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.754.8
Subject(s) - quail , biology , transgene , yolk sac , embryonic stem cell , embryogenesis , embryo , microbiology and biotechnology , ex vivo , andrology , germline , in vivo , gene , genetics , endocrinology , medicine
The purpose of current study is to establish a transgenic Japanese quail model for studying the involvement of genes in regulating nutrient utilization during embryonic development. A week before hatching is the rapid growth period of avian embryonic development, approximately 68 % of the lipid contents in the yolk are absorbed. However, the mechanism by which the yolk sac membrane (YSM) transports lipids remains unclear. We injected the high titer pLenti‐CAG‐eGFP to Japanese quails embryos and successfully generated 6 sexually mature chimera founders (F0) out of 1000 injection, and produced germ‐line transmission offsprings (both F1 and F2) via mating. The eGFP positive rate ofchorioallantoic membrane (CAM) in hatched F1 and F2 are approximately 15‐20% and 4%. eGFP served as a marker during selection. We confirmed the existence of transgenic DNA in the blood cells or of F1 and F2 birds by RT‐PCR. Tissues are also examined for the expression of transgene by histological analysis. The results suggested that eGFP was ectopic expressed in heart, liver and feather in F1 and F2 transgenic quails. In order to clarify the role of endodermal epithelium cell (EEC) that YSM plays in the process of embryonic development, we established a primary endodermal epithelial cells (EECs) from ED (embryonic day) 5 eggs as ex vivo culture system, and culture in DMEM/F12 medium with 10% calf serum. After getting EECs, most tissues could sccessfully generate ex vivo culture cells. These systems can be used to determine the involvement of individual genes during embryonic development.