Gene expression profiles of tissues and cell-lines can be powerful tools for
documenting the genetic response to a particular treatment, such as stressors.
However, there is a paucity of information on the genetic stress response in the
brain. Therefore, we attempted to profile gene expression in the brain of juvenile
steelhead trout (Oncorhynchus mykiss) in response to stressors commonly
encountered in aquaculture settings and similar to those encountered in hydropower
dam mitigation efforts.
We subjected fish to a combined out-of-water and low-water stressor
totaling three hours. Plasma stress response factors indicate that fish were
undergoing a physiological stress response after 3 hours of continuous stressor. We
utilized suppression subtractive hybridization to identify cDNA fragments up- or
down-regulated in the brain upon completion of the stressor. Forward and reverse
subtractions, and sub-cloning of the purified PCR products yielded 59 clones all of
which were sequenced. Sequenced cDNA fragments were subjected to BLASTn
and BLASTx searches over the course of one year. Fragments fell into the
following functional categories: those associated with ATP generation, signal
transduction, ion transport, translational machinery, DNA packaging and
mobilization, cell structure, and cDNA fragments with cryptic function. Of the 59,
12 were selected for further analysis, and 5 were confirmed to be differentially
expressed by northern hybridization. The differentially expressed genes included
cytochrome b, NADH dehydrogenase subunit 2, ATPsynthase subunit 6, a cDNA
fragment with unknown function, and neuron specific gene 1.
Our results present a first attempt to profile gene expression in the brain of
fish and demonstrate the power of molecular tools at capturing large amounts of
biological information without having to target any one particular gene. A gene
expression profile of the brain consequent to stress provides a catalog of responses
at a given time point. This catalog can then be used to isolate full-length cDNAs,
localize mRNAs in the brain or other tissue, as probes to determine expression
patterns and time courses of gene expression in other tissues, and for the
quantification of cDNA molecules with real time PCR. / Graduation date: 2003
| Identifer | oai:union.ndltd.org:ORGSU/oai:ir.library.oregonstate.edu:1957/30350 |
| Date | 03 December 2002 |
| Creators | Schwindt, Adam R. |
| Contributors | Schreck, Carl B. |
| Source Sets | Oregon State University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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