Author
|
Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal Publication Acceptance Date: 1/8/2014 Publication Date: 2/18/2014 Citation: Liu, S., Gao, G., Palti, Y., Cleveland, B.M., Weber, G.M., Rexroad III, C.E. 2014. RNA-seq analysis of early hepatic response to handling and confinement stress in rainbow trout. PLoS One. 9(2). e88492. DOI: 10.1371/journal.pone.0088492. Interpretive Summary: Aquaculture is the fastest-growing animal food producing sector of agriculture, and rainbow trout is one of the most important aquaculture species as it is cultured on every continent except Antarctica. Fish under intensive rearing conditions experience various stressful conditions such as handling, crowding, sub-optimal water quality and temperature fluctuations. Stress has been shown to have negative impacts on survival, growth, reproduction and fillet quality, therefore to improve production efficiency it is crucial to understand stress responses at the physiological and molecular levels. To this end we sequenced high volumes of RNA from fish subjected to handling and confinement to identify 316 genes which play roles in responding to stress, providing a better understanding of the physiological response. Technical Abstract: Fish under intensive rearing conditions experience various stressors which have negative impacts on survival, growth and fillet quality. Identifying and characterizing the molecular mechanisms underlying stress responses will facilitate the development of strategies that aim to improve animal welfare and aquaculture production efficiency. In this study, we used RNA-seq to identify transcripts which are differentially expressed in the rainbow trout liver in response to handling and confinement stress due which are relevant to aquaculture production. Total RNA was extracted from the livers of individual fish in five tanks having eight fish each, including three tanks of fish subjected to handling and confinement stress and two control tanks. Equal amount of total RNA of six individual fish was pooled by tank to create five RNA-seq libraries which were sequenced in one lane of Illumina HiSeq 2000. Three sequencing runs were conducted to obtain a total of 491,570,566 reads which were mapped onto the previously generated stress reference transcriptome to identify 316 differentially expressed transcripts (DETs). Twenty one DETs were selected for qPCR to validate the RNA-seq approach. The fold changes in gene expression identified by RNA-seq and qPCR were highly correlated (r = 0.94). Several gene ontology terms including transcription factor activity and biological process such as glucose metabolic process were enriched among these DETs. Pathways involved in response to handling and confinement stress were implicated by mapping the DETs to reference pathways in the KEGG database. |
