發明
中華民國
100100352
I 412590
一種嵌合性內部核醣體進入區序列及其用途
中原大學
2013/10/21
本實驗根據Rhopalosiphum padi virus (RhPV) 5’ IRES上6個TAAG桿狀病毒晚期及最晚期啟動子之特徵序列所在的位置進行序列刪除,發現RhPV 5’ IRES隱藏的晚期啟動子,主要的活性決定範圍位於全長579個核苷酸的第309~418之間,其含有兩個相連的TAAG motif且啟動子活性達全長的60%。我們將該段序列命名為RP110 啟動子。接著我們藉由融合腸病毒71型的IRES片段與RP110組合來提高RP110的轉譯活性。此一組合序列即為一段於桿狀病毒表現系統與哺乳類表現系統同時兼具功能的人工基因工程元件。 Series of deletion experiments were conducted in six baculovirus late promoter conservative motifs (TAAG) in the cDNA of RhPV 5’ IRES to determine the specific motif which provide cryptic promoter activity of RhPV 5’ IRES. Two tandem repeated TAAG motifs within 309 ~ 418 nts of the 579 nts was responsible for critical promoter activity in baculovirus infected Sf21 cells and the sequence segment was named RP110. Thus, chimeric sequence was formed by fusing RP110 with cDNA derived 5’UTR of enterovirus 71 to generate an artificial sequence . In conclusion, the chimeric sequence will facilitate the development of a dual-baculovirus shuttle expression vector that can express genes in insect cells as well as mammalian cells.
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