RNAi Silencing of Exogenous and Endogenous Reporter Genes Using a Macrocyclic Octaamine as a "Compact" siRNA Carrier. Studies on the Nonsilenced Residual Activity
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- 作者：Kazuki Matsui ; Shohei Horiuchi ; Shinsuke Sando ; Takashi Sera ; and Yasuhiro Aoyama
- 刊名：Bioconjugate Chemistry
- 出版年：2006
- 出版时间：January 2006
- 年：2006
- 卷：17
- 期：1
- 页码：132 - 138
- 全文大小：312K
- 年卷期：v.17,no.1(January 2006)
- ISSN：1520-4812
文摘
A macrocyclic octaamine 1 having a covalently linked lipid-bundle structure was introduced as a new type ofsiRNA carrier. Gel electrophoresis, DLS, and SPR results indicate that it strongly binds to a luciferase-targeting21-mer (42P) siRNA with a ratio of 1/P 
0.3 (1/N 2.4) to give remarkably compact 1-siRNA complexeswith an average size of ~10 nm. The 1-mediated siRNA silencing of the exogenous luciferase gene occurs witha 90-95% efficiency. The overall suppression-[siRNA] profile with a 5-10% residual activity in the saturationregion is commonly observed irrespective of the cell type (HeLa, HepG2, or HEK293), the order, or timing(stepwise or simultaneous) of supply of the siRNA and that of the luciferase-encoding plasmid, the level ofmRNA transcribed, or the type of carriers (1 vs lipofectamine 2000). The silencing of the endogenous DsRed2gene stably incorporated in the genome of HeLa cells also has a similar overall profile. These results suggest that(1) the cellular uptake of the plasmid and that of the siRNA are basically independent of each other and (2) theincomplete silencing is not due to insufficient siRNA delivery. Implication of item 2 is briefly discussed.
0.3 (1/N 2.4) to give remarkably compact 1-siRNA complexeswith an average size of ~10 nm. The 1-mediated siRNA silencing of the exogenous luciferase gene occurs witha 90-95% efficiency. The overall suppression-[siRNA] profile with a 5-10% residual activity in the saturationregion is commonly observed irrespective of the cell type (HeLa, HepG2, or HEK293), the order, or timing(stepwise or simultaneous) of supply of the siRNA and that of the luciferase-encoding plasmid, the level ofmRNA transcribed, or the type of carriers (1 vs lipofectamine 2000). The silencing of the endogenous DsRed2gene stably incorporated in the genome of HeLa cells also has a similar overall profile. These results suggest that(1) the cellular uptake of the plasmid and that of the siRNA are basically independent of each other and (2) theincomplete silencing is not due to insufficient siRNA delivery. Implication of item 2 is briefly discussed.