LUND, S.G.*; WHITMER, A.C.; HOFMANN, G.E.; Univ. of California, Santa Barbara; Univ. of California, Santa Barbara; Univ. of California, Santa Barbara: A PCR-based approach to examining the loss of inducible heat-shock protein expression in an Antarctic fish.

The heat shock proteins (Hsps) are an example of a group of proteins which have been evolutionarily conserved over the broadest range of taxa. Therefore, evidence of disruption in the pathway of inducible Hsps in an organism, such as is the case in some Antarctic notothenioid fishes, provides an opportunity for investigating the function and evolution of protein production at the molecular level. This project is using both DNA- and RNA-based approaches to look for changes in nucleic acid sequence and gene copy number in differentially adapted notothenioid fish. Two primer sets were used to amplify Hsp70 for DNA probe development: previously developed trout-specific primers and primers designed from highly conserved gene regions. Probe specificity was confirmed against samples from fish with demonstrated Hsp induction under thermal stress. Probes were used for Southern blot analysis of hsp gene copy number in 3 fish species: the focus Antarctic species, Trematomus bernacchii, which does not show a predicted heat shock response under experimental conditions; Bovichtus variegates, a closely related, cold-temperate fish from New Zealand that has the predicted induction of Hsps under experimental heat shock conditions; and Notothenia angustata, another cold-temperate fish that may be an intermediary species. Primers were also used on genomic DNA from the same three fish species to examine both intra- and inter-specific DNA sequence variation. Further applications of primers and probes will be discussed. Supported by NSERC PDF to SGL and NSF grant OPP 0087971 to GEH.