Project description:Rainbow smelt (Osmerus mordax) are freeze-resistant fish that accumulate glycerol and produce an antifreeze protein during winter. Quantitative reverse transcription-PCR (qPCR) and subtractive hybridization studies have revealed a few genes in smelt liver to be differentially regulated in winter in comparison with the fall when water temperatures are warmer. In order to further define the suite of processes that are regulated seasonally, we undertook a large-scale analysis of gene expression by hybridization of smelt cDNA to the salmonid 16K cGRASP cDNA microarray. These microarray experiments were conducted as a focused sieving exercise, which identified informative genes for further study in the microarray samples and over a seasonal sampling series using quantitative reverse-transcription PCR.
Project description:Rainbow smelt (Osmerus mordax) are freeze-resistant fish that accumulate glycerol and produce an antifreeze protein during winter. Quantitative reverse transcription-PCR (qPCR) and subtractive hybridization studies have revealed a few genes in smelt liver to be differentially regulated in winter in comparison with the fall when water temperatures are warmer. In order to further define the suite of processes that are regulated seasonally, we undertook a large-scale analysis of gene expression by hybridization of smelt cDNA to the salmonid 16K cGRASP cDNA microarray. These microarray experiments were conducted as a focused sieving exercise, which identified informative genes for further study in the microarray samples and over a seasonal sampling series using quantitative reverse-transcription PCR. Total RNA was obtained from livers of 3 male fish sampled on October 20th and aliquots were pooled to contain equimolar concentrations of each RNA preparation. Total RNA was prepared from 3 male fish on January 3rd and processed in the same manner. Fluorescently labeled cDNA preparations were made using the fall (October) and winter (January) pools and mixed before hybridization to the microarray. The cDNAs were hybridized to three separate microarrays in order to generate technical replicates. Differential expression was assessed in order to select genes for further study.