TY - JOUR
T1 - Two-dimensional single strand conformation polymorphism (SSCP) of 16S rRNA gene fragments reveals highly dissimilar bacterial communities in an acidic fen
AU - Schmalenberger, Achim
AU - Tebbe, Christoph C.
AU - Kertesz, Michael A.
AU - Drake, Harold L.
AU - Küsel, Kirsten
PY - 2008/9
Y1 - 2008/9
N2 - Genetic fingerprinting methods such as denaturing gradient gel electrophoresis (DGGE) and single strand conformation polymorphism (SSCP) are only able to separate about 20-40 well-distinguishable bands (signals) within each sample. As a result, the diversity of 16S rRNA genes within biological samples may be underestimated, because multiple sequences can migrate at the same rate to form a single band. This study reports a two-dimensional SSCP fingerprinting method that has the capability to resolve hundreds of signals in a single fingerprint by using different gel temperatures in the two dimensions of the separation (20 °C and 30 °C, respectively). Unlike previous two-dimensional approaches, the method presented in this study does not rely on DNA products of variable lengths but is able to separate 16S rRNA gene fragments of the same length. To demonstrate the effectiveness of this new method, DNA samples from oxic and anoxic zones of an acidic fen were examined. Whereas one-dimensional SSCP fingerprints indicated high similarity (>93%) between 16S rRNA gene fragments from oxic and anoxic zones of the fen, the two-dimensional SSCP approach virtually found no similarities.
AB - Genetic fingerprinting methods such as denaturing gradient gel electrophoresis (DGGE) and single strand conformation polymorphism (SSCP) are only able to separate about 20-40 well-distinguishable bands (signals) within each sample. As a result, the diversity of 16S rRNA genes within biological samples may be underestimated, because multiple sequences can migrate at the same rate to form a single band. This study reports a two-dimensional SSCP fingerprinting method that has the capability to resolve hundreds of signals in a single fingerprint by using different gel temperatures in the two dimensions of the separation (20 °C and 30 °C, respectively). Unlike previous two-dimensional approaches, the method presented in this study does not rely on DNA products of variable lengths but is able to separate 16S rRNA gene fragments of the same length. To demonstrate the effectiveness of this new method, DNA samples from oxic and anoxic zones of an acidic fen were examined. Whereas one-dimensional SSCP fingerprints indicated high similarity (>93%) between 16S rRNA gene fragments from oxic and anoxic zones of the fen, the two-dimensional SSCP approach virtually found no similarities.
KW - Bacterial diversity
KW - Fen
KW - Two-dimensional SSCP
UR - http://www.scopus.com/inward/record.url?scp=56049111212&partnerID=8YFLogxK
U2 - 10.1016/j.ejsobi.2008.07.002
DO - 10.1016/j.ejsobi.2008.07.002
M3 - Article
AN - SCOPUS:56049111212
SN - 1164-5563
VL - 44
SP - 495
EP - 500
JO - European Journal of Soil Biology
JF - European Journal of Soil Biology
IS - 5-6
ER -