SignalSilence? Atg9A siRNA I

• 产品编号：CST-7051S      品牌：CST       原厂货号：7051S
• 产品分类：DNA和RNA纯化 > RNA干扰 > RNAi Oligos > siRNA
• 应用分类：

描述：

SignalSilence® Atg9A siRNA I 能抑制人、小鼠、大鼠和猴Atg9A表达。来自Cell Signaling Technology (CST)的SignalSilence^® Atg9A siRNA I 可以帮助研究者通过RNA干扰特异性地抑制 Atg9A 的表达，这种方法可以通过将双链RNA分子传递到细胞内从而使基因表达有选择的沉默。来自CST的所有的SignalSilence^® siRNA产品都是经过内部严格检测的，并且通过Western blot 分析证明确实能够减少目的蛋白的表达。通过三苯甲基分析每个碱基以监测寡核苷酸的合成，确保合适的配对效率。随后寡核苷酸通过亲和固相萃取法纯化。退火的RNA双链通过质谱分析来证实其精确的组成。每一批产品都通过质谱分析与前面的产品进行比较，来保证不同批次之间的最大一致性。CST推荐使用100 nM SignalSilence^® Atg9A siRNA I 进行转染，48到72小时后对细胞进行裂解。转染步骤按照转染试剂说明书提供的步骤进行。遇到任何使用方面的问题，请随时联系CST。每小瓶可供100次转染，每次转染量相当于在转染24孔板时，每孔总体积为300μl培养基中siRNA的终浓度为100nM。

自噬是自噬溶酶体对其包裹的细胞质内含物进行降解的一种分解代谢过程(1,2)。自噬过程一般是在营养匮乏的条件下被激活，但是也与一些生理过程有关，如发育、分化、神经变性、感染和癌症(3)。在酵母中已阐明自噬的分子机制，这是由许多自噬相关(Atg)基因调控(4)。

Atg9，一个在酵母中发现的Atg蛋白，对于自噬体的形成至关重要(5)。有两种基于酵母同源 Atg9p的人功能性同源基因：一种是Atg9A，也被确认为Atg9L1 和mAtg9；另外一种是Atg9L2，它被首次确认为氮氧合成酶3反义物(NOS3AS)(6,7)。Atg9A是一个完整的膜蛋白，是启动自噬体形成和膨大所必须的(6，7)。自噬体膜招募Atg9A 是一个动态且短暂的过程，同时，Atg9A也在自噬相关结构高尔基体转运网络(TGN)和核内体之间周期循环，在自噬体膜上形成一个非稳定的结构(6，8)。Atg9A转运的精确调控机制尚未完全阐明，但研究表明其涉及p38促分裂原活化蛋白激酶 (MAPK)-结合蛋白p38IP和Beclin-1结合蛋白Bif-1(9,10)。

1. Reggiori, F. and Klionsky, D.J. (2002) Eukaryot Cell 1, 11-21.
2. Codogno, P. and Meijer, A.J. (2005) Cell Death Differ 12 Suppl 2, 1509-18.
3. Levine, B. and Yuan, J. (2005) J Clin Invest 115, 2679-88.
4. Klionsky, D.J. et al. (2003) Dev Cell 5, 539-45.
5. Noda, T. et al. (2000) J Cell Biol 148, 465-80.
6. Young, A.R. et al. (2006) J Cell Sci 119, 3888-900.
7. Yamada, T. et al. (2005) J Biol Chem 280, 18283-90.
8. Orsi, A. et al. (2012) Mol Biol Cell 23, 1860-73.
9. Webber, J.L. and Tooze, S.A. (2010) EMBO J 29, 27-40.
10. Takahashi, Y. et al. (2011) Autophagy 7, 61-73.

原厂资料：

Homology

Species predicted to react based on 100% sequence homology: Mouse, Rat, Monkey

Specificity / Sensitivity

SignalSilence® Atg9A siRNA I inhibits human, mouse, rat, and monkey Atg9A expression.

Description

SignalSilence® Atg9A siRNA I from Cell Signaling Technology (CST) allows the researcher to specifically inhibit Atg9A expression using RNA interference, a method whereby gene expression can be selectively silenced through the delivery of double stranded RNA molecules into the cell. All SignalSilence® siRNA products from CST are rigorously tested in-house and have been shown to reduce target protein expression by western analysis.

Quality Control

Oligonucleotide synthesis is monitored base by base through trityl analysis to ensure appropriate coupling efficiency. The oligo is subsequently purified by affinity-solid phase extraction. The annealed RNA duplex is further analyzed by mass spectrometry to verify the exact composition of the duplex. Each lot is compared to the previous lot by mass spectrometry to ensure maximum lot-to-lot consistency.

Directions for Use

CST recommends transfection with 100 nM SignalSilence® Atg9A siRNA I 48 to 72 hours prior to cell lysis. For transfection procedure, follow protocol provided by the transfection reagent manufacturer. Please feel free to contact CST with any questions on use. Each vial contains the equivalent of 100 transfections, which corresponds to a final siRNA concentration of 100 nM per transfection in a 24-well plate with a total volume of 300 μl per well.

Background

Autophagy is a catabolic process for the autophagosomic-lysosomal degradation of bulk cytoplasmic contents (1,2). It is generally activated by conditions of nutrient deprivation but is also associated with a number of physiological processes including development, differentiation, neurodegeneration, infection, and cancer (3). The molecular machinery of autophagy was largely discovered in yeast and is directed by a number of autophagy-related (Atg) genes (4). 
 Atg9, one of the Atg proteins identified in yeast, is essential for autophagosome formation (5). There are two human functional orthologues based on the yeast homolog Atg9p: Atg9A, which has also been identified as Atg9L1 and mAtg9, and Atg9L2, which was first reported as nitric-oxide synthase 3 antisense (NOS3AS) (6,7). Atg9A is an integral membrane protein that is required for both the initiation and the expansion of the autophagosome (6,7). Recruitment of Atg9A to the autophagosomal membrane is dynamic and transient as Atg9A also cycles between autophagy-related structures known as omegasomes, the trans-Golgi network (TGN), and endosomes, and at no point becomes a stable component of the autophagosomal membrane (6,8). The precise regulation of Atg9A trafficking is not fully clarified, yet it is suggested to involve p38 mitogen-activated protein kinase (MAPK)-binding protein p38IP and the Beclin-1-binding protein Bif-1 (9,10).

1. Reggiori, F. and Klionsky, D.J. (2002) Eukaryot Cell 1, 11-21.
2. Codogno, P. and Meijer, A.J. (2005) Cell Death Differ 12 Suppl 2, 1509-18.
3. Levine, B. and Yuan, J. (2005) J Clin Invest 115, 2679-88.
4. Klionsky, D.J. et al. (2003) Dev Cell 5, 539-45.
5. Noda, T. et al. (2000) J Cell Biol 148, 465-80.
6. Young, A.R. et al. (2006) J Cell Sci 119, 3888-900.
7. Yamada, T. et al. (2005) J Biol Chem 280, 18283-90.
8. Orsi, A. et al. (2012) Mol Biol Cell 23, 1860-73.
9. Webber, J.L. and Tooze, S.A. (2010) EMBO J 29, 27-40.
10. Takahashi, Y. et al. (2011) Autophagy 7, 61-73.

注意事项：

Storage: Atg9A siRNA I is supplied in RNAse-free water. Aliquot
and store at -20ºC.