2.4.3. DGGE analysis of PCR products

We used the phorU electrophoresis system (INGENY, Leiden, The Netherlands) for conducting DGGE (Fischer & Lerman, 1983), applying a specifically designed protocol for running ciliate PCR products. The PCR products were analyzed on a 15% (w/v) polyacrylamide gel, with a denaturing gradient from 5% to 60% (100% denaturant correspond to 7 mol/l urea and 40% formamide). Due to the high polyacrylamide concentration, the casted polyacrylamide gel mix was allowed to polymerize for 4 hr. Prior to loading samples onto the gel, the PCR products were mixed with loading buffer (40% [w/v] glycerol, 60% [w/v] 1× Tris‐acetate‐EDTA [TAE], bromphenol blue) at a sample/buffer ratio of 1:4. The gel was run in 1× TAE (40 mmol/l Tris, 20 mmol/l acetate, 1 mmol/l EDTA) at a constant voltage of 100 V for 24 hr and at a temperature of 60°C. After gel electrophoresis, the gel was stained in a 1× SybrGold solution (Invitrogen™ S11494, Thermo Fisher Scientific GmbH, Dreieich, Germany) for 50 min, followed by destaining in distilled water for 5 min. Finally, the gel was visualized under UV light. Differences in band migration distance allowed the identification of individual strains (see Appendix B for details).