Question 1

使用KinaseMax 5’末端标记试剂盒时我的磷酸化DNA不能连接。我应该怎么办？

Accepted Answer

我们建议使用酚/氯仿提取加乙醇沉淀的方法纯化激酶反应产物，然后洗脱DNA。如果这样仍然不行，可以使用离心柱层析或变性聚丙烯酰胺凝胶电泳进一步纯化反应产物。

Question 2

我的样本在使用KinaseMax 5’末端标记试剂盒时没能很好地被标记，但是阳性对照标记正常。可能的原因是什么？

Accepted Answer

核酸底物中可能有抑制剂。如有必要，可以使用酚：氯仿：异戊醇提取加离心柱纯化的方法纯化您想要标记的核酸。另外，RNA的5’末端有可能因为三级结构不容易被接触到。在这种情况下，可以尝试在反应前将RNA在90°C加热5分钟，然后立刻将其置于冰上。

Question 3

对于非同位素标记探针和放射性同位素标记探针您们分别提供哪些探针制备试剂盒？

Accepted Answer

我们提供两种非同位素探针标记试剂盒：

•The BrightStar Psoralen-Biotin Nonisotopic Labeling Kit (Cat. No. AM1480)
•MAXIscript Kits for in vitro transcription (Cat. Nos. AM1308 through AM1326)

我们提供3种放射性同位素探针标记试剂盒：

- 用于体外转录的MAXIscript试剂盒（货号AM1308至AM1326）
- KinaseMax5’末端标记试剂盒（货号AM1520）
- DECAprime II DNA标记试剂盒（货号AM1455）

Question 4

My phosophorylated DNA won't ligate when using the KinaseMax 5'; End-Labeling Kit. What can I do?

Accepted Answer

We recommend cleaning up the kinase reaction using a phenol/chloroform extraction followed by ethanol precipitation, then eluting the DNA. If this doesn't help, further purify the reaction by spin-column chromatography or denaturing polyacrylamide electrophoresis.

Question 5

My samples are not labeling well when using the KinaseMax 5' End-Labeling Kit, but the positive control works. What could be the problem?

Accepted Answer

There may be an inhibitor in the nucleic acid substrate. If needed, clean up the nucleic acid you want to label with a phenol: chloroform: isoamyl alcohol extraction followed by spin-column purification. Alternatively, the 5' end of the RNA may be inaccessible because of tertiary structure. In this case, try heating the RNA to 90&#176;C for 5 minutes, then place it immediately on ice just before the reaction.

Question 6

What probe kits do you offer for nonisotopic probe labeling and radiolabeled probe labeling?

Accepted Answer

We offer two nonisotopic probe labeling kits:

- The BrightStar Psoralen-Biotin Nonisotopic Labeling Kit (Cat. No. AM1480)
- MAXIscript Kits for in vitro transcription (Cat. Nos. AM1308 through AM1326)

We offer three radiolabeled probe labeling kits:

- MAXIscript Kits for in vitro transcription (Cat. Nos. AM1308 through AM1326)
- KinaseMax 5' End-Labeling Kit (Cat. No. AM1520)
- DECAprime II DNA Labeling Kit (Cat. No. AM1455)

KinaseMax™ 5' End-Labeling Kit - FAQs