BioProject

Vertebrates are colonized at birth by complex microbial communities (microbiota) that influence diverse aspects of host biology. More...

Vertebrates are colonized at birth by complex microbial communities (microbiota) that influence diverse aspects of host biology. We have used a functional genomics approach to identify zebrafish genes that are differentially expressed in response to the microbiota. We assessed RNA expression profiles from zebrafish larvae at 6 days post-fertilization (dpf) that were either raised continuously in the absence of any microorganism (germ-free or GF), or raised GF through 3dpf then colonized with a normal zebrafish microbiota (conventionalized or CONVD). Overall design: Total RNA was purified from pooled intact zebrafish larvae (28-80 larvae/pool, 3 biological replicate pools/condition) using Trizol reagent (Invitrogen) followed by DNase I digestion (DNA-Free, Ambion) according to manufacturers' protocols. Total RNA from each replicate pool (12ug RNA/replicate) was used as template for independent cDNA synthesis and in vitro transcription reactions (BioArray HighYield RNA Transcript Labeling Kit; Enzo Life Sciences) to generate biotinylated cRNA targets. cRNA targets (20ug/replicate) were fragmented using standard methods. Hybridization and scanning were performed using standard Affymetrix protocol. Raw expression values were normalized (Invariant set method) and modeled (PM-MM model), and present/absent calls were generated using dChip software (build date Dec.11, 2005). Less...