| Olfactory is important to perceive the outside world, and play a role in search of food,exchange in groups, locate geolocation and other acts. The realization of the function of the olfactory is based on the implementation of the olfactory receptor protein function. The OR is able to identify and combine odor molecules in the outside world. This is the starting point of olfactory function. After a long evolution, OR having five independent family, one of them is the main olfactory receptor. In quantity, it is the most than others, and it can identify and combine with the sodor molecules that dissolve in water. in study, C. nasus MOR-4K13 and MOR-51i2 gene belong to the MOR. By prediction, OR has seven transmembrane structure with α-helix, and is a member of G protein coupled receptor(GPCRs) family. Reported from the research that the olfactory functions play a role in the fish migration behavior. By research, C. nasus has two kinds of ecotype: migration and settlement. the migratory behavior for spawning migratory.This study analysed C. nasus OR gene cloning, gene structure, bioinformatics analysis,expression differences of organization, system evolution and the differences in amino acid sequence and relationship of system evolutionary in different ecotype OR gene. For subsequent research such as factors influencing C. nasus migratory behavior, "olfactory orientation hypothesis" and "olfactory imprint theory" provides useful data.1. Cloning and tissues expression of C. nasus olfactory receptor gene MOR-4K13. Using rapid amplification of cDNA ends, the MOR-4K13 gene were received from migratory C. nasus in Jingjiang river. This gene was 1098 bp in length.the open reading frame which encoded 320 amino acid residues was 963 bp in length and is located in 51~1013bp, and the gene was single exon. The protein had seven hydrophobic and alpha helix transmembrane structure, belonged to the G-protein-coupled receptor superfamily A, and had two natural ligands: cholesterol andoleic acid. There were no signal peptide structure, belonged to non-secreted protein, and did not contain disulfide bond structure. This protein could phosphorylate by 22 serine,27 threonine, 17 tyrosine, and had 8 protein binding region, 5 DNA-binding region and5 RNA-binding region. The MOR-4K13 protein was part of plasma membrane components. Partial structure was intrinsic component of membrane, and other partial structure was integral. This the protein had the activity of transmembrane signaling receptor, GPCRs, signal transducer, molecular transducer, olfactory receptor. And had sensory perception of chemical stimulus, signal transdution, cell communication regulation of biological process, cellular response to stimulus, regulation of cellular process, multicellular organismal process and biological regulation. The homology in MOR-4K13 gene of C. nasus and other fishes which had been reported was 65% to84%. The MOR-4K13 protein was 40% to 68%, which the homology of closely related species C. harengus was 77% and 68%. In the evolutionary tree, recently, evolutionary distance in C. nasus and C. harengus were close, both in the same small branches. In evolutionary distance, both and Salmoniformes, Cyprinodontiformes, Lepisosteiformes,Tetraodontiformes, Pleuronectiformes were distant. Based on qRT-PCR among 10 tissues and organs of the settlement population in the poyang lake, the result revealed that the MOR-4K13 gene was expressed strongly in olfactory rosette and gonad than others. The weak expression quantity expressed in muscle,eye, stomach, liver and gills,but almost not in the heart. In addition, the MOR-4K13 gene overall expression quantity in migratory C. nasus olfactory rosette in Jingjiang river above the sedentary in Poyang lake. In summary, the MOR-4K13 gene not only was related to olfactory function, but also participated in gonadal development or associated with reproductive migration,existed in gender gap in the olfactory ability.2. Cloning and tissues expression of C. nasus olfactory receptor gene MOR-51I2. Using genome walking technique, the MOR-51I2 gene were received from migratory C. nasus in Jingjiang river. The open reading frame which encoded OR protein was 999 bp in length,and the gene was single exon. The 3 untranslated regionsof this gene has microsatellite sequence that is formed from(AC)n repeating units that there are several T and G, and have the different length in different geographical populations. The MOR-51I2 gene could code 332 amino acid residues that also had seven hydrophobic and alpha helix transmembrane structure, belonged to the G-protein-coupled receptor superfamily A. There were no signal peptide structure and did not contain disulfide bond structure. This protein could phosphorylate by 22 serine,25 threonine, 17 tyrosine, and had 9 protein binding region, 6 DNA-binding region and7 RNA-binding region.The MOR-51I2 protein was part of plasma membrane components. Partial structure was intrinsic component of membrane, and other partial structure was integral. This the protein had the activity of transmembrane signaling receptor, GPCRs, signal transducer, molecular transducer, olfactory receptor. The homology in MOR-51I2 gene of C. nasus and other fishes which had been reported was66% to 85%. The MOR-51I2 protein was >51%, which the homology of closely related species C. harengus was 77% and 68%. In the evolutionary tree, recently, evolutionary distance in C. nasus and C. harengus were close, both in the same small branches.Based on quantitative real-time PCR among 10 tissues in the settlement population of Poyang lake, the result showed that the MOR-51I2 gene was expressed strongly in olfactory rosettes and gonad than others. The MOR-51I2 gene was almost not express in the stomach, liver, gills, muscle, heart or eye. The expression quantity in female olfactory rosettes was 2 times higher than that in male olfactory rosettes in the migratory population, the MOR-51I2 gene overall expression quantity in migratory C.nasus olfactory rosette in Jingjiang river above the sedentary in Poyang lake.From the above, MOR-51I2 gene not only was related to olfactory function,but also took part in gonad development or breeding migratory, and involved in the differentiation of ecotype. |
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