eGFP is a commonly used reporter gene. eGFP mRNA directly expresses protein in the cytoplasm without relying on activation. The protein expression speed is faster than transfected DNA, and there is no risk of genome integration. Protein expression is directly related to the amount of mRNA. Cells transfected with eGFP mRNA can express strong and bright green fluorescent protein, and the excitation/emission wavelengths are 488 nm/509 nm, respectively. eGFP mRNA can be used as a standard to detect the transfection efficiency of different transfection reagents, and can also be used as a control to study the transfection and expression of fluorescent proteins in mammalian cells.
By simulating the mRNA processing process in eukaryotes, the product has a Cap 1 cap structure at the 5' end and a poly(A) tail at the 3' end, which increases the stability and translation efficiency of mRNA[1]. N1-methyl-pseudouridine (1-methylpseudouridine, m1ψ) is a methyl modification of naturally occurring pseudouridine, formed by N1-specific pseudouridine A that exists in archaea and eukaryotes It is catalyzed by the base transferase Nepl[2]. N1-methyl-pseudouridine is a substitute for uridine (U), which can effectively enhance RNA stability while reducing anti-RNA immune response [3].
References:
[1]. Jemielity J, Fowler T, Zuberek J, et al. Novel "anti-reverse" cap analogs with superior translational properties. RNA. 2003;9(9):1108-1122
[2]. Callum J C Parr, et al.N 1-Methylpseudouridine substitution enhances the performance of synthetic mRNA switches in cells. 2020 Apr 6;48(6):e35. doi: 10.1093/nar/gkaa070.
[3]. Pedro Morais, Hironori Adachi, Yi-Tao Yu.The Critical Contribution of Pseudouridine to mRNA COVID-19 Vaccines. 2021 Nov 4;9:789427. doi: 10.3389/fcell.2021.789427.
绿色荧光蛋白eGFP是一种常用于分子生物学研究的荧光报告基因。eGFP mRNA能够在不依赖于启动子的情况下直接在细胞质中表达蛋白,蛋白表达速度比转染DNA更快,蛋白表达量与mRNA的转染量直接相关,并且没有基因整合的风险。eGFP mRNA转染细胞后能够表达强烈且明亮的绿色荧光蛋白eGFP,激发/发射光波长分别为488 nm/509 nm。eGFP mRNA能够作为标准品检测不同转染试剂的转染效率,也能够作为对照研究哺乳动物细胞中荧光蛋白的转染和表达。
通过模拟真核生物中mRNA加工过程,该产品的5'端具有Cap 1帽结构,3'端具有poly(A)尾,增加了mRNA的稳定性和翻译效率[1]。N1-Me-pUTP是天然存在的假尿苷pUTP的甲基修饰物,由存在于古细菌和真核生物中的N1特异性假尿苷甲基转移酶Nepl催化生成[2]。该产品使用N1-Me-pUTP替代UTP,有效增强了RNA稳定性,同时降低抗RNA免疫应答[3]。