Molecular cloning, tissue distribution and the expression in the regulation of food intake of prepro-orexin in Nile tilapia ( Oreochromis niloticus  )

We cloned the full length of tilapia prepro-orexin cDNA using RT-PCR and rapid-amplification of cDNA ends (RACE). The full-length of prepro-orexin cDNA was 648 bp containing an open reading frame of 423 bp. The 140 amino acid prepro-orexin protein included a 37 AA signal peptide, a 43 AA Orexin-A and, and 28 AA Orexin-B and the end of the 32 AA peptide of unknown function. The expression of prepro-orexin on tissue distribution, peri-prandial changes, starvation and re-feeding were quantified by real-time PCR. We found that prepro-orexin mRNA was present in all tissues tested and that the highest level was observed in hypothalamus. Expression levels were significantly higher at mealtime (0 h) than before (-2 h, -1 h) and after (+1 h, +2 h) mealtime. Fasting for 2, 4, 6 and 8 d caused significant increases in prepro-orexin mRNA expression in the hypothalamus, and after re-feeding, expression levels of prepro-o rexin mRNA returned to the same level compared to that in the fed group.

Keywords
Nile tilapia (Oreochromis niloticus); Orexin; Cloning; Expression; Food intake