探討聚乙烯亞胺轉殖綠螢光蛋白基因進入老鼠胚胎纖維母細胞的最佳條件

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Title:	探討聚乙烯亞胺轉殖綠螢光蛋白基因進入老鼠胚胎纖維母細胞的最佳條件 Efficacy of polyethylenimine mediated EGFP gene into mouse embryonic fibroblasts
Authors:	張萬豐 Wan-Feng Chang
Contributors:	莊一全嘉南藥理科技大學：生物科技研究所
Keywords:	細胞毒性轉殖率基因轉殖聚乙烯亞胺;老鼠胚胎纖維母細胞;N/P率 cytotoxicity transfection rate gene transfection polyethylenimine Mouse Embryonic Fibroblasts N/P rate
Date:	2008
Issue Date:	2008-12-29 15:19:53 (UTC+8)
Abstract:	聚乙烯亞胺 (polyethylenimine，PEI) 是一個帶有高正電荷密度的有機大分子，每三個原子有一個可質子化的氨基氮。PEI有直鏈跟網狀聚合形式，根據文獻報導不論直鏈或網狀聚合的PEI都顯示有特定的基因轉殖能力。由於PEI劑量往往跟轉殖率成正比，但使用過多時會導致細胞死亡，劑量過少又導致基因轉殖率不高，因此影響其轉殖率的條件將在本研究中被詳加探討。PEI可以和DNA形成一個複合物，而PEI/DNA的複合物可因帶正電的關係，和帶負電的細胞表面吸附，經由胞飲作用進入細胞中，再藉由質子海綿脹破的機制，使得PEI/DNA複合物釋放到細胞質中，並避免了溶酶體的分解，進而將DNA傳送到細胞核，因此達成基因殖入細胞而表現特定的蛋白質的目的。我們調控不同的分子量的PEI及N/P率，對老鼠胚胎纖維母細胞(MEF)做基因轉殖效率及細胞毒性的探討。瞭解PEI的分子量及N/P率對MEF轉染效果的關聯性，並且更深入研究pH值對PEI轉染基因的能力是否能更進一步的加強。研究結果顯示，在最佳的條件下基因轉殖MEF，使用25K L-PEI轉殖率為34%，而25K B-PEI轉殖率為29%。 Polyethylenimine (PEI) is the organic macromolecule with the highest cationic-charge-density potential. Every third atom is an amino nitrogen that can be protonated. Acid-catalyzed polymerization of aziridine produces a highly branched network that can ensnare DNA. PEI retains a substantial buffering capacity at virtually any pH. The polymer comes in two forms: linear and branched. Both of them showed successful gene delivery. The dose of PEI influences transfection rate in gene delivery. Overdose makes cytotoxicity, but low dose of PEI is insufficient for high transfection rate. Understanding how molecular weight and N/P rate of the PEI connected with transfection, and further study in whether decrease the pH enhance the performance on gene transfection. Besides, we probe into how different molecular weight of PEI affect the performance of gene transfection. The subject is to increase the efficiency of gene delivery by PEI and eliminate the cytotoxicity to cell. These finding demonstrate that under our optimal condition, 25K L-PEI and 25K B-PEI have 34% and 29% transfection efficient.
Relation:	校內外完全公開
Appears in Collections:	[Dept. of Biotechnology (including master's program)] Dissertations and Theses

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