CDNA Cloning, Localization, Tissues Expression And Molecular Evolution Of Preprohrelin In Duck, Goose And Emu

Ghrelin is an endogenous ligand for growth hormone seretagogue receptor (GHS-Rs). This 28-amino acid peptide possesses a unique serine residue at the third position (Ser3) that is modified by n-octanoic acid. Ghrelin regulates pituitary growth hormone (GH) secretion .In the study ,we obtained the cDNA sequences of preproghrelin and the cDNA encoding the ghrelin of duck,goose and emu by using RT-PCR and 3' -RACE methods.and then deduced the amino acid sequences of preproghrelin and ghrelin in duck.goose and emu. Sense and antisense primers were designed on the basis of chicken ghrelin cDNA sequence (dEST accession no. AB075215, 836bp) and proventriculus cDNA was performed as the template. By comparing the CDS and amino acid sequences it indicate ghrelin may had little change during the evolution of poultry. The results of phylogenetic analysis showed that the amino acid sequences of duck preproghrelin and goose preproghrelin clustered together firstly .and then came thoses of emu .chicken and Meleagris gallopavo preproghrelin, and then came those of Rana catesbeiana,Oreochromis mossambicus and Oreochromis niloticus , amino acid sequences of preproghrelin of Bos Taurus and Ovis aries came subsequently. It indicated that the preproghrelin of duck,goose and emu had a close phylogenetic affinity to poultry preproghrelin sequences. The clustering results of amino acid sequences of preproghrelin accorded with morphological classification system.On the basis of chicken preproghrelin DNA sequence (accession no AY303688.) we designed sense and antisense primers to screen the chicken BAC library of our lab, consequently we got the BAC clone containing the interned sequence and used it as the probe .Then by FISH method we localized chicken preproghrelin at GGA2q.l .1.According to chicken GAPDH cDNA sequence (dEST accession no. K01458, 1284bp), we designed the primers and regard the PCR productions as the internal standard.Then by using RT-PCR analysis, high levels of expression of emu ghrelin mRNA were detectable in proventriculus were detected, low levels of expression of duck ghrelin mRNA were also detectable in lung muscle ileum duodenum corpus striatum cerebellum brain stem and gizzard.