Observation of bias associated with re-amplification of DNA isolated from denaturing gradient gels

• Published in: FEMS microbiology letters 2005-03, Vol.244 (2), p.385-390
• Main Authors: Nikolausz, Marcell, Sipos, Rita, Révész, Sára, Székely, Anna, Márialigeti, Károly
• Format: Article
• Language: English
• Subjects: 16S rDNA; Action of physical and chemical agents on bacteria; Amplification; Bacteriology; Bias; Biological and medical sciences; Deoxyribonucleic acid; DGGE; DNA; DNA, Ribosomal - analysis; DNA, Ribosomal - genetics; Electrophoresis; Electrophoresis, Polyacrylamide Gel - methods; Fundamental and applied biological sciences. Psychology; Gel electrophoresis; Gels; Microbial activity; Microbiology; Microorganisms; Mycological methods and techniques used in mycology; Mycology; Nucleic Acid Denaturation; PCR; Polymerase chain reaction; Polymerase Chain Reaction - methods; Re-amplification; Soil Microbiology
• ISSN: 0378-1097; 1574-6968; 1574-6968
• DOI: 10.1016/j.femsle.2005.02.013

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.