Multi-generation E. coli-C. elegans Interactions

E. coli HB101, AZ121 and GS035 strains were grown in LB media containing no antibiotics, ampicillin (100 µg/ml) or chloramphenicol (25 µg/ml) respectively (OD595 = 0.6–0.8). Cultures were centrifuged and the re-suspended pellets were seeded onto NGM plates. For one generation feeding experiments, animals were placed on the seeded NGM plates and allowed to grow until young adult stage. For three generation feeding experiments, L1 larval stage animals were placed on the seeded NGM plates and allowed to grow for three generations. Total RNA from harvested animals was isolated using the TriSure reagent (Bioline). Small RNA sequencing libraries of C. elegans were prepared using the NEBNext Multiplex Small RNA Library Prep set (NEB) and the small RNA sequencing libraries of E. coli are prepared using the TruSeq Small RNA set (Illumina). Libraries were either sequenced on a MiSeq or a HiSeq 1500 machine (Illumina). Data analysis was performed as previously described21 (link). Small RNA-seq data is available through NCBI-GEO with accession number GSE64990. qRT-PCR is performed with primers described in Liu et al17 (link). OxyS RNA structure is determined using the Mfold web server22 (link).