T7 RNA Transcription Enzyme Mix (C09-1077-335)

Product DescriptionAs a biological macromolecule, mRNA can only be synthesized on a large scale by in vitro transcription (IVT). T7 promoter is one of the promoters with the highest transcription efficiency at present. Therefore, more synthetic products can be obtained by in vitro transcription with T7 RNA polymerase. T7 RNA transcription enzyme mix optimized the transcription reaction system, synthesized RNA complementary to one strand of DNA from the downstream of template DNA T7 promoter, and simply and quickly obtained a large number of RNA molecules. One reaction can be transcribed up to 150-200 μ G RNA, the transcribed synthetic RNA can be used for downstream applications such as mRNA vaccine preparation, RNA structure and function research, RNase protection, probe hybridization, RNAi, microinjection and in vitro translation.T7 RNA transcription enzyme mix is expressed by large-scale fermentation of E. coli, produced with raw and auxiliary materials of pharmaceutical specifications, and strictly controlled the host protein residue and nucleic acid residue. The product production and quality management procedures in line with GMP specifications ensure that the production process and all raw and auxiliary materials can be traced.Quality requirements project  standard appearance  Clear liquid Visible foreign matter  Compliance with regulations PH value  7.5-8.5 performance  1μL Enzyme mix products can be transcribed no less than 100 μgRNA Endonuclease residues  Degradation of substrate shall not exceed 10% Exonuclease residues  Degradation of substrate shall not exceed 10% RNase residue  Degradation of substrate shall not exceed 10% Bacterial endotoxin content  ≤10EU/ml Exogenous DNA residue  ≤100pg/mg Host protein residue  ≤50ppm Mycoplasma detection  negative Heavy metal residues  ≤10ppm Follow the following specifications1. ISO 9001:2015, certified facility。2. GMP appendix - cell therapy products State Drug Administration.3. general introduction to human gene therapy - Chinese Pharmacopoeia 2020, National Pharmacopoeia Committee.4. USP chapter <1043>, adjuvant materials for cell, gene, and tissue engineered products.5. USP chapter <92>, growth factors and cytokines used in cell therapy manufacturing.6. Ph. Eur.  General chapter 5.2.12, raw materials of biological origin for the production of cell-based and gene therapy medical products.Product usage1) Synthesize single-stranded RNA for mRNA vaccine preparation;2) Synthesize highly specific RNA probes;3) Synthesize siRNA precursor;4) Precursor for RNA splicing reaction (RNA splicing).Storage temperature-20±5 ℃。Matters needing attention1. template efficiency and incubation time:This kit uses 1 μ The template input of G can generate 150-200 μ However, the yield of different templates will vary depending on the sequence, structure, length, purity of the template and the sequence and length of a specific RNA polymerase promoter. Pollutants that affect transcription yield include ribonuclease or pollutants, such as phenol, trace metals and SDS.2. optimized response:The recommended reaction conditions can be applied to the in vitro transcription of most templates. However, for some templates, the yield can be improved by prolonging the reaction time (4 hours overnight reaction) and increasing the amount of templates.3. formwork content:The following table summarizes our experience in regulating the number of templates. The results may vary depending on the template used. Extending the reaction time to 4-6 hours can increase RNA production. Template quantity  RNA production 1000ng(1μg) 130-160μg 500ng(0.5μg) 110-130μg 100ng(0.1μg) 30-50μg 50ng(0.05μg) 15-25μg 10ng(0.01μg) 10-20μg 1ng(0.001μg) 3-8μg 4. maintain RNase free environment:Use RNase free tubes and pipettes;Gloves should be worn when handling kit components or samples containing RNA, and gloves should be changed frequently, especially after contacting potential pollution sources of RNase, such as door handles, pens, pencils and human skin.When not in use, all reagents shall be well sealed. During incubation, all tubes containing RNA were sealed.5. due to 10 × The concentration of the transcription buffer is high, and the high salt environment will lead to the inactivation of the polymerase. At the same time, the buffer contains spermidine, which will form precipitation with the template DNA. When preparing the reaction solution, it is necessary to adjust the sampling sequence of the components, calculate the system, add water first, then buffer, NTP, and finally add template and enzyme to prevent 10 × The high concentration of salt ions affected the enzyme. Specifications and Purity: Pharmaceutical grade. Related Document: https://ald-pub-files.oss-cn-shanghai.aliyuncs.com/aladdinsci/pdp/sds/1/T489304-SCI_d99a5816bf187df2f754389731d336d5.pdf.