Observation of bias associated with re-amplification of DNA isolated from denaturing gradient gels.
DNA from environmental PCR products separated by denaturing gradient gel electrophoresis (DGGE) was isolated from the background smear rather than from discrete bands of the DGGE gel. The "interband" region was considered as a potential source of less dominant members of natural microbial communities. Surprisingly, instead of detecting new bands from the re-amplified PCR products, patterns very similar to the original ones were obtained regardless of the position of the "interband" region. The results suggest that the separation of amplicons by DGGE may not be perfect and band re-amplification based sequence analyses need careful interpretation.
Nikolausz
Marcell
M
Department of Microbiology, Eötvös Loránd University of Science, Pázmány Péter sétány 1/c, 1117 Budapest, Hungary. marcell.nikolausz@ufz.de
Sipos
Rita
R
Révész
Sára
S
Székely
Anna
A
Márialigeti
Károly
K
D004275:DNA, Ribosomal / Q000032:analysis* / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D009691:Nucleic Acid Denaturation; D016133:Polymerase Chain Reaction / Q000379:methods*; D012988:Soil Microbiology
D004275:DNA, Ribosomal