| 查看: 2211 | 回复: 1 | |||
[求助]
RNA与镁离子
|
|
求助下面这段描述中的镁离子是有什么作用的？另外镁离子与RNA的稳定与构象有关吗？原理是什么呢？ A T7 primer and gene-specific primers were used to PCR amplify pri-miR- 1792 sequences from plasmid DNA template. PCR products were gelpurified and used as templates for in vitro transcription using the Riboprobe (Promega) system together with 32P-CTP for radioactive labeling. Microprocessor purified from Flag-DROSHA-293 cells was used for in vitro Microprocessor assays (Gregory et al., 2004). Cold RNA was produced using the same strategy without 32P-CTP addition. For RNA annealing, 10 mM Sample Text was added to 200 pmol cold RNA and incubated at 95C for 5 min and then slowly cooled to reverse transcription (RT). Annealed RNA was subjected to 5% native polyacrylamide gel for ethidium bromide staining and used for Microprocessor assay followed by small RNA northern blot analysis. His- CPSF3 complex was purified from E. coli as described previously for other proteins (Chang et al., 2013). Assays conditions were as for Microprocessor. |
回复此楼» 猜你喜欢
生物质炭与活化剂氢氧化钾混合注意事项
已经有0人回复
-
科研人必读：新一代 KRAS G12C 抑制剂 Olomorasib 的研究进展
已经有0人回复
-
化学工程及工业化学论文润色/翻译怎么收费?
已经有87人回复
-
BT-11（Omilancor）研究化合物全解析：从分子结构到实验应用要点
已经有0人回复
-
富马酸贝达喹啉：二芳基喹啉类工具化合物的化学特征
已经有0人回复
-
BT投稿with editor状态快两周了，正常吗
已经有25人回复
-
Silevertinib（BDTX-1535）结构拆解：炔基双环片段为何成为合成瓶颈
已经有0人回复
-
RO4929097 解析：双苯并氮杂䓬骨架与 Notch 通路实验设计要点
已经有0人回复
-
分子量 1157 的 PROTAC 怎么配不析出：SGK3-PROTAC1 的溶解、分装与图谱复核
已经有0人回复
-
logP 6.1 的羧酸怎么配母液：GSK 650394 的溶解窗口与稀释顺序
已经有0人回复
-
从"水不溶"到 71 mg/mL：雷特格韦的 pH 依赖溶解与螯合色谱边界
已经有0人回复
|
In Vitro Transcription, Microprocessor, and CPSF3 Cleavage Assays A T7 primer and gene-specific primers were used to PCR amplify pri-miR- 1792 sequences from plasmid DNA template. PCR products were gelpurified and used as templates for in vitro transcription using the Riboprobe (Promega) system together with 32P-CTP for radioactive labeling. Microprocessor purified from Flag-DROSHA-293 cells was used for in vitro Microprocessor assays (Gregory et al., 2004). Cold RNA was produced using the same strategy without 32P-CTP addition. For RNA annealing, 10 mM MgCl2 was added to 200 pmol cold RNA and incubated at 95C for 5 min and then slowly cooled to reverse transcription (RT). Annealed RNA was subjected to 5% native polyacrylamide gel for ethidium bromide staining and used for Microprocessor assay followed by small RNA northern blot analysis. His- CPSF3 complex was purified from E. coli as described previously for other proteins (Chang et al., 2013). Assays conditions were as for Microprocessor. |
2楼2015-09-23 18:37:14
