Most eukaryotic mRNAs require a 7-methyl guanosine (m7G) cap structure at the 5´-end and a poly(A) tail at the 3′-end for efficient translation. The HiScribe T7 mRNA Kit with CleanCap Reagent AG utilizes an optimized RNA synthesis formulation and trinucleotide cap analog technology for co-transcriptionally capping mRNAs that contain a natural Cap-1 structure in a single simplified reaction without compromising RNA yield. By using a DNA template with a T7 promoter sequence followed by an AG initiation sequence and an encoded poly(A) tail, mRNAs can be transcribed with a 5´-m7G Cap-1 structure that is polyadenylated, translationally competent and able to evade the cellular innate immune response.
The HiScribe T7 mRNA Kit with CleanCap Reagent AG is formatted with individual vials of NTPs and CleanCap Reagent AG to allow for partial or complete substitution of modified NTPs, with a total kit yield of 1.8 mg of mRNA. Cap-1 mRNA synthesized from this kit is suitable for many applications, including transfections, microinjections, in vitro translation, preclinical mRNA therapeutic mRNA studies as well as RNA structure and function analysis.
This kit contains sufficient reagents for 20 reactions (20 µl each). Each standard reaction yields ≥ 90 µg of RNA from 1 µg CLuc AG Control Template DNA. Each kit can yield ≥ 1.8 mg RNA.
Figure: 1: CleanCap Reagent AGFIgure 2: Schematic of CleanCap Reagent AG Promoter and Initiation SequenceFigure 3: CleanCap Reagent AG results in higher mRNA synthesis yield than the ARCA analogAll reactions were performed with 5 mM CTP, 5 mM UTP and 6 mM ATP. Standard IVT reactions included 5 mM GTP and no cap analog. ARCA reactions contained a 4:1 ratio of ARCA:GTP (4mM:1mM). IVT with CleanCap Reagent AG contained 5 mM GTP and 4 mM CleanCap Reagent AG and was performed according to recommended protocol (Standard mRNA Synthesis, HiScribe T7 mRNA Kit with CleanCap Reagent AG). Reactions were incubated for 2 hours at 37°C, purified and quantified by NanoDrop®.
This product is related to the following categories:
RNA Synthesis Products
This product can be used in the following applications:
核糖核酸酶A是内切核糖核酸酶，可特异地攻击RNA上嘧啶残基的3'端，切割与相邻核苷酸形成的磷酸二酯键。反应终产物是嘧啶3'磷酸及末端带嘧啶3'磷酸的寡核苷酸。无辅因子及二价阳离子存在时，核糖核酸酶A的作用可被胎盘RNA酶抑制剂(B1ackburn et al.1977)或氧钒—核糖核苷复合物(Puskas et al.1982)所抑制。核糖核酸酶A用途生化研究，测定核酸的结构RNase 保护检测去除非... 查看更多>
