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題名 | 構築於反轉錄病毒載體之豬生長素基因在小鼠胚幹細胞基因組中之嵌插及表現=Integration and Expression of Porcine Growth Hormone Gene after being Transfected Into the Mouse Embryonic Stem Cells by Retroviral Vector |
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作者姓名(中文) | 郭孟富; 陳全木; 吳和光; 鄭登貴; | 書刊名 | 中國畜牧學會會誌 |
卷期 | 27:1 1998.03[民87.03] |
頁次 | 頁59-73 |
分類號 | 437.21 |
關鍵詞 | 小鼠; 胚幹細胞; 反轉錄病毒載體; 基因轉殖; Mouse; Embryonic stem cell; Retroviral vector; Gene transfer; |
語文 | 中文(Chinese) |
中文摘要 | 本研究旨在藉反轉錄病毒載體,將豬生長素因轉染進入小鼠之胚幹細胞,並探討 該外源基因在胚幹細胞內之嵌插及其表現情形。試驗結果顯示,構築於反轉錄病毒載體之豬 生長素基因( pGH-SVDV-Neo, 簡稱 PSN 基因),經 8 次重複感染進入小鼠之同一繼代的 胚幹細胞後,發現該外源基因係以單套且有多處不同嵌插位置之形式,出現於胚幹細胞之基 因組內。 此外,應用北方轉印技術,針對 PSN 基因在胎豬纖維母細胞中與小鼠胚幹細胞內 之轉錄效率進行比較分析,結果證明後者之轉錄效率僅及前者之 4.1 ∼ 38.1% 不等。進一 步針對源自二者萃取獲得基因組 DNA,分別應用具有相同截切位置且對甲基化具敏感性之限 制脢 Hpa II,及對甲基化不具敏感性之限制脢,Msp I,進行截切分析結果,更證明此等嵌 入小鼠胚幹細胞基因組內之 PSN 基因,常有被甲基化之現象; 相類似結果則未見之於其在 胎豬纖母細胞內者。綜合上述結果,應用反轉錄病毒載體確可輕易將豬生長素基因,穩定嵌 插於小鼠胚幹細胞之基因組 DNA 內,但該嵌入之外源基因常因發生甲基化現象, 導致顯著 降低其後續之轉錄效率。 |
英文摘要 | Integration and expression of porcine growth hormone gene in the mouse embryonic stem cell (ESC), after being transfected by means of retrovirus as a gene vector, were studied in the present studies. Experimental results appeared that there were multiple insertions, but each was with a single copy of retrovirus gene porcine GH-SVDV-Neo (PSN) integrated within the genome of mouse ESC, after they had been subjeected to eight times of repeated infections by retrovirus PSN within 48 h of culture period between passage 6 and 7. However, evidence from the Northern blot analysis confirmed that the transcription activity of PSN pene integrated into the mouse ESC was much less than that in the fetal fibroblast cells of porcine (4.1~38.2% vs 100% ). It was found that the retrovirus PSN gene was often methylated after they had been integrated into the mouse ESC while the same phenomenon was not observed in those of fibroblast cells. |
本系統之摘要資訊系依該期刊論文摘要之資訊為主。