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No bands observed in PCR products

No bands observed in PCR products - Agarose Gel Electrophoresis Forum

No bands observed in PCR products - Agarose Gel Electrophoresis Forum. A forum to discuss electrophoresis staining, troubleshooting, and development of films for DNA and RNA gels.


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Old 04-09-2011, 11:42 PM
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Default No bands observed in PCR products



I had 5 PCR products and a ladder run in 2% Agarose gel.
However the UV lamp showed that only ladder had orange color, and not even that had bands going down the gel. (in other words it was orange only in the well that I pippetted in)

As I was disposing the gel, I noticed that my gel seemed thicker than
other people's gels. Is it possible that my wells were too deep and that caused no band observation? Is it likely my PCR samples failed because my master mix was incorrectly made?
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Old 04-11-2011, 06:14 AM
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Post Re: No bands observed in PCR products

Hi,
There may be many reasons for not getting bands. The solution however is that we have to rule out the possibilities one by one.
1. There may be a problem with your gel preparation. As you have mentioned that the orange colour was not seen even in the ladder, it seems your gel is not prepared correctly. So its better if you check the buffer composition and pH and also the Ethidium bromide concentration.
2. You can check whether your gel is prepared perfectly, by just running the ladder alone and comparing the bands with the reference bands (given in the guide)
3. If there is no problem with the gel and still you didn't get the bands, then we have to check with the master mix.
4. Before that check the concentration of template.
There is not so much to do with the depth of the well.

Hope this was useful.
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