PureLink™ 快速凝胶提取试剂盒

以下是为您提供的一些建议：

•许多酶，包括限制性内切酶和连接酶，可被少量琼脂糖和高氯酸盐污染物抑制。正常情况下，这并不会对实验造成影响。但是，如果出现这样的问题，您可使用未经再纯化的DNA，通过增加消化或连接过程中酶的用量或通过延长消化或连接的孵育时间优化实验。另外，也可以在使用相同的反应体系和相同用量的限制性内切酶的情况下，减少消化或连接反应的DNA量。
•洗脱片段中可能有残留乙醇。一定要完全离心以去除洗涤缓冲液并丢弃，使用新试管收集洗脱的DNA。在对乙醇非常敏感的应用中，应将吸附柱打开后，在室温下直立放置15分钟，使多余的乙醇挥发。
•洗涤步骤可能未达到理想效果。在这种情况下，洗脱液中可能含有痕量高氯酸盐。为避免这种情况发生，可将离心时间延长至5分钟，并使用洗涤缓冲液洗2次。
•如果您使用了高浓度琼脂糖或向柱子中加入超过250 mg琼脂糖，应确保执行备选的洗涤步骤（可参考QC protocol）。在对EDTA敏感的应用中，应使用水（pH 7.5–8.5）或无EDTA的10 mM Tris（pH 8.0）洗脱。

常规和低熔点琼脂糖凝胶均适用。当凝胶浓度超过2%时，每100 mg 胶使用600 μL溶胶液进行溶解。

可以，用水洗脱是可以的，不过请确保水质洁净，pH介于7.5和8.5之间。

可以，我们的凝胶提取试剂盒和TAE及TBE琼脂糖凝胶均兼容。

Here are some suggestions for your experiments:

- Many enzymes, including restriction endonucleases and ligases, are inhibited by small amounts of agarose and by perchlorate contaminants. This is not normally a problem. If it is, however, you can use the DNA without repurification by increasing the amount of enzyme used in the digest or ligation or by increasing the digestion incubation time. Also, you may use less DNA in your digest or ligation with the same total volume and the same amount of restriction enzyme.
- There may have been residual ethanol in the eluted fragment. Be sure to thoroughly centrifuge to remove the wash buffer, discard the wash buffer, and use a fresh tube to collect the eluted DNA. For applications that are very sensitive to ethanol, let the open spin column stand for 15 minutes at room temperature to let any excess ethanol evaporate.
- The washing steps may not be as efficient as they should be. Under these circumstances, there may be trace amounts of perchlorate in the eluate. To avoid this, extend the centrifugation times to 5 minutes and wash 2 times with wash buffer.
- Be sure to perform the optional wash step if you are using higher concentrations of agarose or are adding more than 250 mg to the cartridge. If applications are sensitive to EDTA, elute with water (pH 7.5-8.5), or with 10 mM Tris, pH 8.0 without EDTA.