16s rRNA sequencing of stool pellets from IEC-Bmal1-/- mice and Bmal1flox controls in naive and DSS-treated states over the course of a day
16s rRNA sequencing of stool pellets from IEC-Bmal1-/- mice and Bmal1flox controls in naive and DSS-treated states over the course of a day
Fecal pellets were collected and snap frozen. Once thawed, pellets were mixed with PBS and homogenised.  Feel pellets were collected from IEC-Bmal1-/- and Bmal1flox controls in naive state at ZT0, ZT4, ZT8, ZT12, ZT16, ZT20. Mice were then treated with 2% DSS. After 4 days of DSS, feel pellet collection was repeated. Microbial DNA was extracted with the DNeasy PowerSoil Pro Kit (Qiagen). Pre-amplification of the V4 region of 16S rRNA was performed using forward primer 5'-ACACTCTTTCCCTACACGACGCTCTTCCGAT-CTNNNNNGTGCCAGCMGCCGCGGTAA-3' (annealing sites in bold) and reverse primer 5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTGGACTACHVGGGTWTCTAAT-3'. PhiX control v3 library (PhiX) was spiked into samples to balance low base diversity often found in microbiome samples.