Cloning，identification and tissue expression analysis of nanos3 gene in Monopterus albus

【Objective】To clarify the important role of nanos3 gene in development and maintenance of ovary in Monopterus albus，so as to lay the foundation for studies on visualized molecular marker of PGCs，germ cell transplantation and gonadal differentiation mechanism.【Method】 cDNA sequence of nanos3 gene was cloned by RACE.ProtParam，SOMPA，SWISS-MODEL，SignalP，TMHMM，NetPhos and other online software were used to conduct bioinformatics analysis of nanos3 protein.Expression of nanos3 gene in different tissues of M.albus was determined through quantitative real-time PCR（qRT-PCR）and frozen section fluorescence in situ hybridization method was used to detect cellular location of nanos3 in gonad tissue.【Result】Full length of cDNA sequence of nanos3 gene was 1289 bp，including 5'-UTR of 147 bp，open reading frame（ORF）of 531 bp and 3'-UTR of 611 bp，and 176 amino acid residues were encoded.Molecular weight of nanos3 protein in M.albus was 19.61 kD，with a oretical isoelectric point（pI）of 9.07，indicating it was alkaline protein.In the 108-162th position of amino acid sequence existed 2 zinc finger motif“CCHC”domains，without any signal peptide and transmembrane region.In the secondary structure of the protein，random coil（64.20%）accounted for the highest proportion，followed by α-helix（24.43%），and extension chain and β-turn accounted for 6.82% and 4.55%，respectively.nanos3 amino acid sequence similarity between M.albus and Mastacembelus armatus，Toxotes jaculatrix and Danio rerio was 67.5%，57.2% and 50.3%；similarity between M.albus and amphibian species of Rana chensinensis and Rana rugosa was 39.3% and 30.3%；similarity between M.albus and mammal species of humans and mice was 33.3% and 30.9%，respectively.nanos3 gene was not expressed in the muscle of M.albus，and its expression was low in testis，kidney，spleen，brain，liver，intestine，heart and blood and its expression in ovary was the highest，significantly higher than that in the tissues（P<0.01），showing tissue-specific expression patterns.Fluorescence in situ hybridization（FISH）analysis results of frozen sections showed that nanos3 gene in M.albus was only expressed in germ cells，not in somatic cells.【Conclusion】nanos3 gene evolutes conservatively，and it was mainly expressed in the germ cells of ovary，and mainly expressed in the cytoplasm，and this is related to the transmission of nuclear genetic information.nanos3 geneis the functional gene related to gonadal development and differentiation in M.albus.