I'm trying to amplify rRNA 16S from soil samples using universal bacterial primers F341 with GC clamp and R518 for a DGGE assay.
My problem is I always get two bands, an expected 220bp one and a second of about 300bp.
Did anyone have the same problem? How did you solve it?
Thank you!
Hi Laura,
You may found some answers to your question from a previous similar question posted on RG, here is the link:
https://www.researchgate.net/post/Can_anyone_help_me_with_DGGE_primers
Come back here if this post is not helping you and give more information related to your PCR and gel.
Best,
Aimeric
Hello, Laura
Sometimes the second band in DGGE can be due to the lack of time in the incubation step of PCR, according to Janse et al (2005). I recommend you to read the paper
Ingmar Janse, Jasper Bok, Gabriel Zwart (2004). A simple remedy against artifactual double bands in denaturing gradient gel electrophoresis
Great luck,
Martha
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