title: The microbiota of rotifers propagated on different diets
design: Rotifer samples were taken using plankton nets (100 m), then rinsed with sterile distilled water and stored in microcentrifuge tubes with 2 ml RNAlater solution (Ambion). Water samples (approx. 250 ml) were taken simultaneously from all tanks and immediately filtered through 200 m and 50 m plankton nets in order to remove macro-particles and finally vacuum-filtered using the sterile, disposable all-in-one system (UltraCruz Filter Flask, pore size 0.22 m). The sterile filter papers with bacteria from the tank water were placed in tubes with 15 ml RNAlater solution. Total DNA was extracted from rotifers (pooled wheel animals) samples and filtered water samples stored on RNAlater using the PureLink Genomic DNA Mini Kit (Invitrogen) according to the manufacturer's instructions and partial 16S rRNA gene amplicons that covered the hypervariable regions V1-V3 were obtained with slightly modified universal primers 27FMod and 519RMod. PCR was performed using the HotStarTaq Plus Master Mix Kit (Qiagen). Sequencing of PCR products was performed by Molecular Research LP (TX, USA) on an Ion S5 XL system following the manufacturers guidelines.