產(chǎn)品描述：The High Yield T7 RNA Synthesis Kit與傳統的體外轉錄反應相比，有著(zhù)很高的合成量，能夠多達25倍，每個(gè)反應可以合成高達180μg的RNA。
試劑盒中提供的原料：包括50個(gè)反應，20μL/反應。
| Component | 50 rxn kits | Storage |
| 5× Reaction Buffer* | 200μL | -20℃ |
| 100mM ATP Solution | 100μL | -20℃ |
| 100mM GTP Solution | 100μL | -20℃ |
| 100mM UTP Solution | 100μL | -20℃ |
| 100mM CTP Solution | 100μL | -20℃ |
| Enzyme Mix | 75μL | -20℃ |
| Control Template(0.5μg/μL) | 10μL | -20℃ |
| DNase I (RNase-free, 1U/μL) | 50μL | -20℃ |
| Nuclease-free H2O | 1mL | -20℃ |
| Ammonium Acetate Stop Solution | 1.5mL | -20℃ |
| Lithium Chloride Precipitation Solution | 1.5mL | -20℃ |
| Gel Loading Buffer | 0.5mL | -20℃ |
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注：5× Reaction Buffer儲存在-70℃，可能會(huì )產(chǎn)生白色沉淀，如果有白色沉淀產(chǎn)生，將試劑管加熱至37℃ 5min 并且充分混合使其溶解后使用。
1. 按下表中的量完成20μL反應，可根據需要按照比例放大或者減小
| Component | Amount |
| Nuclease-free H2O | to 20μL |
| 100mM ATP Solution | 2μL |
| 100mM GTP Solution | 2μL |
| 100mM CTP solution | 2μL |
| 100mM UTP solution | 2μL |
| Template DNA | 1μg* |
| 5× Reaction Buffer | 4μL |
| Enzynme Mix | 1.5μL |
*轉錄產(chǎn)量取決于模板的添加量。
2. 充分混合，37℃反應 2h。
3.（可選擇）加入1μL DNase I（RNase-free），充分混合，37℃反應30min。
儲藏溫度：-20℃
質(zhì)控檢測：All components are tested in a functional assay as described in this procedure. A 20-μL reaction containing 1 μg of the control template DNA which codes for a ~800b transcript synthesized >150 μg of RNA after a 2 hr incubation.
1. Milligan JF, Groebe DR, Witherell GW, and Uhlenbeck OC (1987) Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA template. Nucl. Acids Res. 15: 8783–8798.
2. Molecular Cloning, A Laboratory Manual, 2nd edition. (1989) editor C Nolan, Cold Spring Harbor Laboratory Press.