Molecular Cloning And Expression Of Pod1Gene In Medaka (Oryzias Latipes)

Sex determination in animal is one of the most remarkable phenomena in nature, especially in the life sciences research. Podl gene is a member of bHLH transcription factor family, and is involved in regulation of neurogenesis, myogenesis, sex determination and sexual differentiation. However, the function of Podl gene in sex determination and sexual differentiation was not studied in non-mammalian vertebrates. Because medaka is a best model organisms in sex determination and sex differentiation mechanism, we used medaka as a model animal and got results as follows.1.The open reading frame of Podl gene in medaka, which contains516nucleotides coding a protein with171amino acids was cloned. Partial sequences of3’end Podl gene were cloned from medaka testis using rapid amplification of cDNA ends (RACE) procedure.2. Podl gene sequence of medaka was compared to that of other Vertebrate and construct the phylogenetic tree of Podl by bioinformatics software. Synteny map analysis suggest that the Podl gene of medaka was resided the centromeric ragion of medaka linkage group24while that of human was resided the centromeric ragion of human linkage group6the genome. 3. Total RNA was extracted from each organization of medaka, reverse transcribed into cDNA, and then amplified by PCR to detect Podl distribution of gene expression in different tissues of adult medaka. The result showed that the podl gene was exppressed not only in gonad but also in the tissue of heart, knidy and liver, and so on.4.The Podl protein was expressed by the prokaryotic expression vector to predict their protein interaction。In this study, we use medaka as a model animal.we cloned the ORF sequence and the3’end of the gene sequence Podl,and analyzed tissue expression profiles of Podl gene and constructed prokaryotic expression vector. It is make foundation to further study of the molecular mechanism of Podl gene during fish embryo devement and sexual differentation.