| 查看: 1255 | 回复: 5 | ||||
OBM2013铜虫 (小有名气)
|
[求助]
求RNA上某一碱基甲基化的检验方法？
|
| 除了用同位素放射检验16SrRNA上某一碱基甲基化与否外，还有没有别的方法比较简便的验证16SrRNA上的某一碱基是否发生甲基化？求各位大侠赐教！！ |
回复此楼» 收录本帖的淘贴专辑推荐
 [表观遗传学---新四大碱基] | RNA甲基化 |
» 猜你喜欢
高斯计算参数设置
已经有1人回复
-
武汉轻工大学2024年化环学院机械力化学课题组硕士招生
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160人回复
-
负载率计算
已经有0人回复
-
球磨纳米粒子条件
已经有6人回复
-
球磨分散剂
已经有3人回复
-
阴阳离子交换剂！！！！
已经有3人回复
» 本主题相关价值贴推荐，对您同样有帮助:
- zymo 甲基化 求助同仁
已经有15人回复
- 【求助/交流】什么碱基修饰不影响配对但不能够扩增
已经有5人回复
- 避免RNA酶污染的方法
已经有10人回复
OBM2013
铜虫 (小有名气)
- 应助: 0 (幼儿园)
- 金币: 610.2
- 散金: 25
- 帖子: 83
- 在线: 59.4小时
- 虫号: 2347255
- 注册: 2013-03-14
- 性别: GG
- 专业: 兽医免疫学
2楼2013-11-03 09:31:23
deng神之刺客
木虫 (小有名气)
- 应助: 64 (初中生)
- 金币: 2284.9
- 红花: 3
- 帖子: 186
- 在线: 123.8小时
- 虫号: 2747727
- 注册: 2013-10-23
- 性别: GG
- 专业: 植物生理与生化
【答案】应助回帖
★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★ ★
OBM2013: 金币+5, ★有帮助, 谢谢 2013-11-20 11:16:51
OBM2013: 金币+15, ★有帮助 2014-01-08 19:46:06
OBM2013: 金币+5, ★有帮助, 谢谢 2013-11-20 11:16:51
OBM2013: 金币+15, ★有帮助 2014-01-08 19:46:06
| 根据不同检测方法的原理及其发展先后 ,大致可分为三类: (1)将RNA降解为单核苷酸而后采用化学分析仪器如液相色谱或质谱对甲基化与非甲基化的胞嘧啶进行区分; (2)用序列特异性甲基化内切酶使RNA 部份降解 ,通过Nouthern印迹区分甲基化与非甲基化的相关序列;(3)利用亚硫酸氢钠(BISULFITE)预处理 RNA,直接通过引物延伸反应或耦联其他机制对甲基化与非甲基化的胞嘧啶进行区分 |
3楼2013-11-03 10:58:39
OBM2013
铜虫 (小有名气)
- 应助: 0 (幼儿园)
- 金币: 610.2
- 散金: 25
- 帖子: 83
- 在线: 59.4小时
- 虫号: 2347255
- 注册: 2013-03-14
- 性别: GG
- 专业: 兽医免疫学
4楼2013-11-05 12:57:54
ADTechnology 
捐助贵宾 (著名写手)
表观遗传学专业供应商
- 应助: 12 (小学生)
- 金币: 8776.7
- 散金: 1687
- 红花: 1
- 帖子: 1932
- 在线: 42.7小时
- 虫号: 1787705
- 注册: 2012-04-30
- 专业: 人类遗传学
【答案】应助回帖
商家已经主动声明此回帖可能含有宣传内容
★ ★ ★ ★ ★
OBM2013: 金币+5, ★有帮助, 谢谢 2013-11-20 11:12:39
OBM2013: 金币+5, ★有帮助, 谢谢 2013-11-20 11:12:39
|
您可以采用比色法或是qMS-PCR的方法实现，如下是采用比色的方法来实现的。 m6A RNA甲基化定量检测试剂盒(比色法) 货号：A-P-9005 EpiQuik m6A RNA Methylation Quantification Kit (Colorimetric) 背景资料：N6-methyl-adenosine (m6A) is the most common and abundant modification in RNA molecules present in eukaryotes. The m6A modification is catalyzed by a methyltransferase complex METTL3 and removed by the recently discovered m6A RNA demethylases FTO and ALKBH5, which catalyze m6A demethylation in an α-ketoglutarate (α-KG)- and Fe2+-dependent manner. It was shown that METTL3, FTO, and ALKBH5 play important roles in many biological processes, ranging from development and metabolism to fertility. m6A accounts for more than 80% of all RNA base methylations and exists in various species. m6A is mainly distributed in mRNA and also occurs in non-coding RNA such as tRNA, rRNA, and snRNA. The relative abundance of m6A in mRNA transcripts has been shown to affect RNA metabolism processes such as splicing, nuclear export, translation ability and stability, and RNA transcription. Abnormal m6A methylation levels induced by defects in m6A RNA methylase and demethylase could lead to dysfunction of RNA and diseases. For example, abnormally low levels of m6A in target mRNAs due to increased FTO activity in patients with FTO mutations, through an as-yet undefined pathway, contributes to the onset of obesity and related diseases. The dynamic and reversible chemical m6A modification in RNA may also serve as a novel epigenetic marker of profound biological significance. Therefore, more useful information for a better understanding of m6A RNA methylation levels and distribution on RNA transcripts could benefit diagnostics and therapeutics of disease. 产品描述：In this assay, total RNA is bound to strip wells using a RNA high binding solution. m6A is detected using specific capture and detection antibodies. The detected signal is enhanced and then quantified colorimetrically by reading the absorbance in a microplate spectrophotometer at a wavelength of 450 nm. The amount of m6A is proportional to the OD intensity measured. Both negative and positive RNA controls are provided in this kit. A standard curve can be performed (range: 0.02 to 1 ng of m6A) or a single quantity of m6A can be used as a positive control. Because m6A content can vary from tissue to tissue, and from normal and diseased states, or vary under treated and untreated conditions, it is advised to run replicate samples to ensure that the signal generated is validated. This kit will allow the user to quantify an absolute amount of m6A and determine the relative m6A RNA methylation states of two different RNA samples。 m6A RNA甲基化定量检测试剂盒(比色法)步骤示意图 m6A RNA甲基化定量检测试剂盒(比色法)标准曲线 产品特点： 1·Colorimetric assay with easy-to-follow steps for convenience and speed. The entire procedure can be completed within 3 hours and 45 minutes ； 2·High sensitivity, of which the detection limit can be as low as 10 pg of m6A； 3·Unique binding solution allows RNA >70 nts to be tightly bound to the wells, which enables quantification of m6A from both mRNA and ncRNA such as tRNA, rRNA, and snRNA； 4.Optimized antibody and enhancer solutions allow high specificity to m6A, with no cross-reactivity to unmethylated adenosine within the indicated concentration range of the sample RNA； 5.Universal positive and negative controls are included, which are suitable for quantifying m6A from any species； 6.Strip-well microplate format makes the assay flexible for manual or high throughput analysis； 7.Simple, reliable, and consistent assay conditions。 保存建议：在接收到艾德寄出的试剂盒后请按照说明书建议，使用不同的保存条件或温度来保存试剂盒内组分 更多的资料，可以加Q1951545998  |
5楼2013-11-17 12:36:53
6楼2019-05-19 22:32:36
100