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題名 | 經體外成熟和體外受精之牛卵母細胞與卵丘細胞共培養之發育率=Development of Bovine Oocytes Matured, Fertilized and Co-Cultured with Cumulus Cells in Vitro |
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作者 | 李善男; 劉振發; 許義明; Lee, Shan-nan; Lieu, Chen-fa; Hsu, Yi-ming; |
期刊 | 中國畜牧學會會誌 |
出版日期 | 19971200 |
卷期 | 26:4 1997.12[民86.12] |
頁次 | 頁429-438 |
分類號 | 437.3 |
語文 | chi |
關鍵詞 | 牛; 卵母細胞; 體外受精; 卵丘細胞; 胚發育; Bovine; Oocyte; In vitro fertilization; Cumulus cell; Embryo development; |
中文摘要 | 本研究之目的在建立牛卵母細胞經體外成熟和體外受精後,與卵丘細胞作共培養 育成囊胚之簡易方式。 自屠宰場取得完整之黃牛、水牛及乳牛卵巢,置於 37 ℃,0.9% 生 理鹽水中,於一小時內帶回實驗處理。 以 18 號針頭吸取直徑 2~5mm 濾泡內之卵。將卵丘 卵母細胞複合體置於成熟液中,進行體外成熟, 並與解凍後己具獲能之精子完成受精作用, 5 小時後取出受精卵,清洗之,然後再置回含卵丘細胞之培養皿中。受精後 48 小時,卵裂 為 4 細胞期以上之胚被留置於此單層卵丘細胞之共培養液迄 14 天。 結果顯示,黃牛、水 牛及乳牛之卵母細胞受精後 48 小時之卵裂率分別為 37.4(34/91) 、 27.5(14/51) 及 16.7%(4/24); 發育成桑椹胚比率為 24.2 (22/91)、5.9(5/51) 及 12.5% (3/24);發育至 囊胚僅有黃牛及乳牛, 各為 18.7(17/91) 及 12.5% (3/24); 而黃牛能發育到孵化囊胚達 15.4% (14/91)。 以 Fisher Exact Test 檢測其獨立性,由卵母細胞發育至 4~8 細胞,品 種間發育率無顯著差異 (P>0.05)。但由 4~8 細胞發育至桑椹期則差異顯著 (P<0.05)。 結 論建議,利用現階段卵丘細胞共培養系統,可以將黃牛及乳牛之卵母細胞進行體外成熟、體 外受精及體外培養,使其發育至囊胚,而水牛胚之體外生產系統仍須進一步改進。 |
英文摘要 | A procedure to obtain the bovine blastocyst in a repeatable maturation, fertilization and culture system in vitro (IVMFC) with frozen-thawed semen was developed. Native yellow cattle (NYC), water buffalo (WB) and Holstein ovaries were obtained from a slaughterhouse and transported to the laboratory in 0.9% saline at 37 ℃ in one hour. All follicles (2~5 mm in diameter) were aspirated with a 18-gauge needle. Grade 1 and 2 cumulus-oocyte complexes (COCs) were cultured in the maturation medium (TCM-199 with 5% FBS) in a CO �� incubator (2% CO �� 38.5 ℃ ) for 28~30 hrs. The frozen-thawed semen was suspended in Brackett Oliphant (BO) medium for sperm capacitation and its concentration was adjusted to 1 × 10 �� cell/ml. After maturation, the oocytes were subsequently introduced into the drops of sperm suspension. Five hours later, the inseminated oocytes were removed and washed and then introduced into the maturatin medium that had been kept with cumulus cells. Forty-eight hrs after insemination, the oocytes were removed and examined for their cleavage stages; four or more cell embryos were further cultured with cumulus cell monolayer in the dish. The cleavage rate after 48 hrs after insemination of NYC, WB and Holstein were 37.4(34/91), 27.5 (14/51) and 16.7% (4/24) respectively. Embryos that had developed to the morula stage after day 6~7 in this co-culture system were 24.2 (22/91), 5.9(3/51) and 12.5% (3/24), respectively. Blastocysts were obtained from NYC and Holstein groups representing 18.7 (17/91) and 12.5% (3/24). Totally, 14 out of 91 NYC oocytes (15.4%) were cultured to hatched blastocyst. Data were analyzed using Fisher Exact Test. Results of this experiment showed a difference in cleavage rates among the NYC, WB and Holstein embryos developed from 4~8 cells to morula stage. In addition, these data demonstrated that embryos from NYC and Holstein produced in this co-culture system with cumulus cell manolayer were of better quality than those from WB. |
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