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PCR Smearing Troubleshooting

PCR Smearing Troubleshooting - PCR - Polymerase Chain Reaction Forum

PCR Smearing Troubleshooting - PCR - Polymerase Chain Reaction Forum. Discuss and ask questions about PCR troubleshooting, PCR protocols and methods, PCR products, and PCR theory.


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  #1  
Old 11-04-2007, 01:48 PM
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Question PCR Smearing Troubleshooting



Dear forum members,
how do you troubleshoot PCR smearing?
I get smearing that runs all throughout from the lane top to bottom but I can see my band.

How can i get only my band and less smearing?
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Old 11-04-2007, 06:10 PM
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Default Re: PCR Smearing Troubleshooting

Smearing can be from rRNA, too much DNA, too much PCR product---for these you can reduce smearing by cutting back on the template DNA and lessening the number of PCR cycles. Another reason can be degraded DNA,RNA which you can improve by using better preserved/processed samples.
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Old 11-06-2007, 01:37 PM
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Thumbs up Re: PCR Smearing Troubleshooting

thanks Danfive
that was very helpful I will post back what was wrong
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Old 10-27-2008, 03:16 PM
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Default Re: PCR Smearing Troubleshooting

you could try adding a new cycle to your PCR thermal profile after your repetitive cycle i.e add 72C ofr 5-10min for final extension. this may allow incomplete extension to be completed. just a suggestion
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