[Specific-PCR and denaturing gradient gel electrophoresis assistant isolation of Thauera spp. from a coking wastewater treatment plant].
OBJECTIVE
We used specific-PCR and denaturing gradient gel electrophoresis (DGGE) to isolate Thauera spp. from a coking wastewater treatment plant.


RESULTS
To isolate Thauera from the denitrifying bioreactor of a coking wastewater treatment, biofilm was inoculated to six different media and cultured them under both aerobic and anaerobic conditions. We then compared the composition of Thauera spp. using Thauera-specific PCR-DGGE method. The media 1/10 NB and MMQ which grew higher diverse Thauera spp. and fewer colonies, were used to isolate Thauera sp. under aerobic condition. The colonies were then screened by Thauera-specific PCR. The purity of the colonies that shown Thauera-specific PCR positive signal was then checked by DGGE. The colonies with multiple species were further streaked on different media. DGGE analysis showed that Thauera in colony Q20 was enriched in medium MMP. The colony was finally purified by streaking on MMP medium for several rounds. The composition of the colonies were tracked by Thauera-specific PCR and DGGE at each step. Finally, three strains were purified, which were identified as Thauera sp. according to their 16S rRNA gene sequences.


CONCLUSIONS
Guiding with specific biomarker, the efficiency and sensitivity of bacteria isolation can be largely improved.
Mao
Yuejian
Y
Key Laboratory of Microbial Metabolism, Ministry of Education, College of Life Science and Biotechnology, Shanghai Jiaotong University, Shanghai 200240. yuenjian@gmail.com
Zhang
Xiaojun
X
Zhang
Baorang
B
Zhao
Liping
L
D015415:Biomarkers; D019149:Bioreactors / Q000382:microbiology; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013045:Species Specificity; D020628:Thauera / Q000235:genetics / Q000302:isolation & purification*; D014865:Waste Disposal, Fluid; D014871:Water Microbiology
D015415:Biomarkers