SignalSilence?DNMT3L siRNA II (Mouse Specific)

• 产品编号：CST-12308S      品牌：CST       原厂货号：12308S
• 产品分类：DNA和RNA纯化 > RNA干扰 > RNAi Oligos > siRNA
• 应用分类：

描述：

来自Cell Signaling Technology （CST）的SignalSilence® DNMT3L siRNA II (Mouse Specific) 可以通过RNA干扰特异性抑制DNMT3L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建议使用100 nM SignalSilence® DNMT3L siRNA II (Mouse Specific)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。哺乳动物细胞中DNA胞嘧啶甲基化是一种可遗传的表观遗传修饰，并且对于基因表达、基因组印记和发育的正确调控都是至关重要的。目前已经确认了3中哺乳动物甲基转移酶家族：DNMT1、DNMT2和 DNMT3 （1,2）。DNMT1在增殖细胞中为组成型表达，是一种维持型甲基转移酶，负责为复制过程中新合成的DNA转移正确的甲基化模式。DNMT3A和DNMT3B 在胚胎干细胞中强烈表达而在成熟的体细胞组织中表达减弱。他们属于从头甲基转移酶，能够甲基化DNA上之前没有被甲基化的区域。DNMT2在成熟体细胞组织中的表达水平较低，它的失活无论是对从头DNA甲基化还是维持型DNA甲基化都没有影响。DNMT3L是DNMT3A 和 DNMT3B从头甲基转移酶的催化失活型调控因子，从而让他们在胚胎干细胞、睾丸、卵巢和胸腺中呈低水平表达（1,2）。这些从头甲基转移酶是一个异源四聚体复合物，包含两分子DNMT3L以及两分子DNMT3A或DNMT3B（3）。DNMT3L包含一个氨基末端ATRX-DNMT3-DNMT3L (ADD)结构域以及羧基端甲基转移酶样结构域（4-7）。甲基转移酶样结构域结合到DNMT3A 和 DNMT3B上通过提高S-腺苷甲硫氨酸和DNA的结合从而促进催化活性（4,5）。ADD结构域能够通过与非甲基化组蛋白H3 Lys4从而将甲基转移酶复合物募集到基因组上转录失活的区域（6,7）。

1. Hermann, A. et al. (2004) Cell Mol Life Sci 61, 2571-87.
2. Turek-Plewa, J. and Jagodziński, P.P. (2005) Cell Mol Biol Lett 10, 631-47.
3. Jia, D. et al. (2007) Nature 449, 248-51.
4. Holz-Schietinger, C. and Reich, N.O. (2010) J Biol Chem 285, 29091-100.
5. Suetake, I. et al. (2004) J Biol Chem 279, 27816-23.
6. Ooi, S.K. et al. (2007) Nature 448, 714-7.
7. Otani, J. et al. (2009) EMBO Rep 10, 1235-41.

原厂资料：

Description

SignalSilence® DNMT3L siRNA II (Mouse Specific) from Cell Signaling Technology (CST) allows the researcher to specifically inhibit DNMT3L 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® DNMT3L siRNA II (Mouse Specific) 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.

ackground

Methylation of DNA at cytosine residues in mammalian cells is a heritable, epigenetic modification that is critical for proper regulation of gene expression, genomic imprinting, and development (1,2). Three families of mammalian DNA methyltransferases have been identified: DNMT1, DNMT2, and DNMT3 (1,2). DNMT1 is constitutively expressed in proliferating cells and functions as a maintenance methyltransferase, transferring proper methylation patterns to newly synthesized DNA during replication. DNMT3A and DNMT3B are strongly expressed in embryonic stem cells with reduced expression in adult somatic tissues. DNMT3A and DNMT3B function as de novo methyltransferases that methylate previously unmethylated regions of DNA. DNMT2 is expressed at low levels in adult somatic tissues and its inactivation affects neither de novo nor maintenance DNA methylation. 
 
 DNMT3L is a catalytically inactive regulatory factor for the DNMT3A and DNMT3B de novo methyltransferases that is expressed at low levels in embryonic stem cells, testis, ovaries, and thymus (1,2). These de novo methyltransferases consist of a heterotetrameric complex containing two molecules of DNMT3L, and either two molecules of DNMT3A or DNMT3B (3). DNMT3L contains an amino-terminal ATRX-DNMT3-DNMT3L (ADD) domain and a carboxy-terminal methyltransferase-like domain (4-7). The methyltransferase-like domain binds to DNMT3A and DNMT3B to stimulate catalytic activity by increasing the binding of S-adenosylmethionine and DNA (4,5). The ADD domain recruits the methyltransferase complex to transcriptionally inactive regions of the genome by binding to unmethylated histone H3 Lys4 (6,7).

1. Hermann, A. et al. (2004) Cell Mol Life Sci 61, 2571-87.
2. Turek-Plewa, J. and Jagodziński, P.P. (2005) Cell Mol Biol Lett 10, 631-47.
3. Jia, D. et al. (2007) Nature 449, 248-51.
4. Holz-Schietinger, C. and Reich, N.O. (2010) J Biol Chem 285, 29091-100.
5. Suetake, I. et al. (2004) J Biol Chem 279, 27816-23.
6. Ooi, S.K. et al. (2007) Nature 448, 714-7.
7. Otani, J. et al. (2009) EMBO Rep 10, 1235-41.

注意事项：

Storage: DNMT3L siRNA II (Mouse Specific) is supplied in RNAse-free water. Aliquot and store at -20ºC.