Rainbow smelt, Osmerus mordax, have an impressive ability to acclimate to very 
cold water. Rainbow smelt exposed to cold (<5 °C) for an extended period of time 
have faster sustained swimming speeds and increased contraction kinetics in 
their myotomal muscle compared to warm acclimated fish. We used RNA Sequencing 
reactions (RNA-Seq) to explore how gene expression underlies thermal acclimation 
by muscle in these fish. Transcriptome analysis is limited in species that lack 
an annotated genome, such as rainbow smelt. The Trinity software package permits 
the de novo assembly of a rainbow smelt transcriptome with a modest learning 
curve. The transcriptome was then analyzed with Kallisto to quantify the 
abundance of each transcript represented in the full transcriptome and Sleuth to 
analyze the resulting RNA-seq datasets. Subsequently qPCR was used to explore 
patterns of thermal acclimation and gene expression for genes of metabolic and 
muscle contractile function. These methodologies revealed shifts in both muscle 
and metabolic gene expression that contribute to the thermal acclimation 
response in rainbow smelt. In fast-twitch, anaerobic white muscle, slow isoforms 
of myosin heavy and light chain tended to be down-regulated with exposure to 
cold in myotomal muscle, while fast isoforms were unchanged. Genes associated 
with protein turnover and aerobic metabolism were up-regulated in the white 
muscle, while those associated with anaerobic metabolism and the cell cycle were 
down-regulated. Collectively the results suggest that thermal acclimation to 
cold is complex process of apparent shifts in gene expression.