Identification of immune-relevant genes from atlantic salmon using suppression subtractive hybridization.
In order to probe the interaction between an invading microorganism and its host, we have investigated differential gene expression in Atlantic salmon (Salmo salar) experimentally infected with the pathogen Aeromonas salmonicida, the causative agent of furunculosis. Subtractive cDNA libraries were constructed by suppression subtractive hybridization (SSH) from 3 immune-relevant tissues at 2 time points during the infection process. Both forward- and reverse-subtracted libraries were generated, and approximately 200 clones were sequenced from each library, giving a total of 1778 expressed sequence tags (ESTs), which were annotated according to functional categories and deposited in GenBank (BQ035314-BQ037059). Numerous genes involved in signal transduction, innate immunity, and other processes have been uncovered in the subtractive libraries. These include known acute-phase reactants, along with more novel genes encoding proteins such as tachylectin, hepcidin, precerebellin-like protein, O-methyltransferase, a putative saxitoxin-binding protein, and others. A subset of genes that were represented in the subtracted libraries was further analyzed by virtual Northern, or reverse transcription-polymerase chain reaction (RT-PCR) assays to verify their differential expression as a result of infection.
Tsoi
S C M
SC
Institute for Marine Biosciences, 1411 Oxford Street, Halifax, Nova Scotia B3H 3Z1, Canada.
Ewart
K V
KV
Penny
S
S
Melville
K
K
Liebscher
R S
RS
Brown
L L
LL
Douglas
S E
SE
D048409:Aeromonas salmonicida; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D020224:Expressed Sequence Tags; D005393:Fish Diseases / Q000276:immunology* / Q000382:microbiology*; D005667:Furunculosis / Q000276:immunology / Q000662:veterinary*; D005786:Gene Expression Regulation / Q000276:immunology*; D015723:Gene Library; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020317:Salmo salar / Q000235:genetics*; D017422:Sequence Analysis, DNA