Cloning, Expression And The Relationship Between DMRT1 Gene And Growth Traits In Pelteobagrus Fulvidraco

DMRT gene family is one of the known control biological development gene families. Its code products with a highly conservative sequences which can combine with the DNA, named DM domain region (the DNA-binding motif shared between files double-sex gene and nematode mab-3 gene). It was able to combine with particular sequences of DNA by the zinc finger structure, adjust the transcription of purpose gene, and control gene expression in gender downstream decision. It plays an important role in the process of sex determination and differentiation. DMRT1 (double-sex and mab3-relatated transcription factorl) is the typical representative of DMRT gene families, which participates widely the sex development of vertebrates and invertebrates, is considered another important gene related to sex-determinating after the discovery of SRY gene and Sox9 gene.To elucidate the role of DMRT1 in the sex determination and differentiation in Pelteobagrus fulvidraco, and provide the theoretical basis for researching of unisexual breeding, growth difference between male and female and sex differentiation mechanism, The full-length DMRT1 cDNA sequences from Pelteobagrus fulvidraco was isolated by RACE SMART and RT-PCR. Its mRNA expression level during the stages of embryonic development, post embryonic development, gonad development and adult was analyzed. Otherwise, the relationship between polymorphism of DMRT1 and growth traits was discussed. The main conclusions are as follows:1. Full-length of DMRT1 cDNA isolation and homology analysisThe full-length of DMRT1 cDNA was 1381 bp, including 30 bp of the 5' untranslated region,454 bp of the 3'untranslated region and 885 bp of the open reading frame, encoding 294 amino acids. Deduced amino acids of Pelteobagrus fulvidraco DMRT1 was compared to Clarias batrachus, Clarias gariepinus and Silurus meridionalis, the similarity were 85%,82% and 85% respectively, while the conserved DM-domain similarity up to 97.2-100%. Other fishes Oreochromis niloticus, Nile tilapia, medaka, zebra fish and rainbow trout shared lower similarity of 60%,60%,53%,64% and 60% respectively, identity of 43% and 45% were also found when compared to mouse and human.2. The level of mRNA expression analysis of DMRT1 geneExpression of DMRT1 in different development stages and adult different tissues of Pelteobagrus fulvidraco were analyzed by RT-PCR and Real-time RT-PCT technologies. The results showed that it expressed throughout the developmental process, expression level of gastrulation period (early, middle) was significantly higher than the rest of the period during embryonic development stage. Expression of the early of post embryonic development (1-35 days after hatching, dah 1-35) was relatively high, its expression reached the highest level at dah 31, and maintained a relatively low level during dah 36-51. DMRT1 mRNA was detected during all development periods of testis with a highest level at periodâ…£, while none throughout ovarian development. In adult male and female, it only expressed in testis. 3. Relationship analysis between DMRT1 polymorphism and the growth traitsUsing PCR-SSCP method analyzed the relationship between DMRT1 polymorphism and growth traits in Pelteobagrus fulvidraco, the results showed that a Gâ†'>A mutation was detected in SS1 site of the extron, which leaded to the amino acid glycine (G) mutation into serine (S). The site was controlled by a pair of alleles (A and B), showing 3 genotypes (AA, BB and AB). Difference analysis between different genotypes and the main growth traits indicated that AA genotype in total length, body length, body height, body thickness and body weight were higher than BB and AB individuals in the female population, in addition to head length, caudal peduncle length and caudal peduncle height. For the male population, in the traits of total length, body height, head length and body weight, AA genotype were the same higher than the BB and AB. But the two analyzed groups did not reach significant level (P>0.05). In the entire population, AA genotype in the full length, body length, caudal peduncle height and body weight were significantly higher than that of BB (P<0.05), extremely significantly higher than AB (P<0.01), significantly higher than BB and AB in body thickness and caudal peduncle length (P<0.05), extremely significantly higher than BB and AB in body height and head length(P<0.01), while BB and AB genotype had no significant differences (P>0.05).The site can be considered as one of candidate markers in the process of marker-assisted breeding in Pelteobagrus fulvidraco, as far as possible increase the AA genotype in the breeding population.