*也被称为 Anti-reverse Cap Analog (ARCA) ,即抗-反向帽类似物。
体外实验的初始阶段,绝大多数真核生物 mRNA 5' 端 m7G 帽结构可促进翻译。对于绝大多数 RNA,帽结构都可提高 RNA 的稳定性,降低其对核酸外切酶降解的敏感性,并促进 mRNA 起始复合体的形成。一些具有 5' 端帽结构的原核 mRNA 也可以像真核 mRNA 一样,能在真核生物无细胞蛋白合成体系中获得高效翻译。另外,还发现在真核生物靶 RNA 的剪切过程同样需要帽结构。
用 7 甲基 G 帽结构类似物,m7G(5' )ppp(5' )G (NEB#S1404) 作为引物能在 SP6 RNA 聚合酶、T7 RNA 聚合酶和 T3 RNA 聚合酶作用下,转录出更多的带帽RNA。因为转录时,T7 RNA 聚合酶掺入的*个碱基是 G,所以用 #S1405 或 #S1406 转录出带有 A-帽结构的 RNA,就不能够翻译,但这对于转染和显微注射实验有很好的稳定性。Contrease 等已经建立了一套方法,以 m7G(5' )ppp(5' )G (NEB #S1404)或 m7G(5' )ppp(5' )A (NEB #S1405) 为引物,利用 E. coli RNA 聚合酶,在体外有效合成带帽 RNA。
帽类似物有两个3' 羟基基团使得它们可从任一方向结合到 RNA 上。而抗-反向帽类似物(ARCA)3' OMethyl-m7G(5' )ppp(5' )G (NEB #S1411) 的一个羟基被甲基化,只能以 m7G 作为*个碱基与 RNA 结合,而这个方向可以增强体外翻译。
N-7003
ARCA (Anti Reverse Cap Analog)
key step in cellular mRNA processing is the addition of a 5’ cap structure, which is a 5'-5' triphosphate linkage between the 5' end of the RNA and a guanosine nucleotide. The cap is methylated enzymatically at the N-7 position of the guanosine to form mature mCAP.
Anderson, B.R., Muramatsu, H., Jha, B.K., Silverman, R.H., Weissman, D., Kariko, K. Nucleoside modifications in RNA limit activation of 2'-5'-oligoadenylate synthetase and increase resistance to cleavage by RNase L (2011) Nucleic Acids Research, EPub Aug | ||||
Warren et al., Highly Efficient Reprogramming to Pluripotency and Directed Differentiation of Human Cells with Synthetic Modified mRNA, Cell Stem Cell (2010), doi:10.1016/j.stem.2010.08.012. | ||||
Anderson, B., Muramatsu, H., Nallagatla, S.R., Bevilacqua, P.C., Sansing, L.H., Weissman, D. & Kariko, K. Incorporation of pseudouridine into mRNA enhances translation by dimishing PKR activation (2010) Nucleic Acids Research, 38(17): 5884-5892. | ||||
Kariko, K., Buckstein, M., Ni, H. & Weissman, D. Suppression of RNA Recognition by Toll-like Receptors: The Impact of Nucleoside Modifiation and the Evolutionary Origin of RNA (2005). Immunity, 23(2), 165-175. | ||||
Peng ZH, Sharma V, Singleton SF, Gershon PD. Synthesis and application of a chain-terminating dinucleotide mRNA cap analog. (2002) Organic Letters. 4(2):161-4. | ||||
Konarska MM, Padgett RA, Sharp PA. Recognition of cap structure in splicing in vitro of mRNA precursors. (1984) Cell. 38(3):731-6. | ||||
Miura K. The cap structure in eukaryotic messenger RNA as a mark of a strand carrying protein information. (1981) Adv Biophys. 14:205-38. | ||||
Banerjee AK. 5'-terminal cap structure in eucaryotic messenger ribonucleic acids. (1980) Microbiol Rev. 44(2):175-205. | ||||
Rosenberg M, Paterson BM. Efficient cap-dependent translation of polycistronic prokaryotic mRNAs is restricted to the first gene in the operon. (1979) Nature. 21;279(5715):696-701. | ||||
Filipowicz W. Functions of the 5,-terminal m7G cap in eukaryotic mRNA. (1978) FEBS Lett. 96(1):1-11. | ||||
Zan-Kowalczewska M, Bretner M, Sierakowska H, Szczesna E, Filipowicz W, Shatkin AJ. Removal of 5'-terminal m7G from eukaryotic mRNAs by potato nucleotide pyrophosphatase and its effect on translation. (1977) Nucleic Acids Res. 4(9):3065-81. |
N-7003-1 | ARCA | 1umole |
N-7003-10 | ARCA | 10umoles |
N-7003-5 | ARCA | 5umoles |