EST analysis and identification of gonad-related genes from the normalized cDNA library of large yellow croaker, Larimichthys crocea

On grounds of the especially limited numbers of identified gonad-specific or gonad-related genes of large yellow croaker Larimichthys crocea which may represent a major obstacle for the study of gonad development and sex differentiation, we initiated a sequencing program of Expressed Sequence Tags (ESTs) in large yellow croaker. In this study, we firstly constructed a normalized gonad cDNA library using the combination of SMART technique and DSN treatment. The titer of amplified cDNA library was 4.8 × 1011 and the percentage of unique cDNA sequences of the library was 82.49%. 2916 unique cDNAs were clustered from the 3535 high quality ESTs. Among the 1785 ESTs which had significant homology with known genes in the NCBI database, about 64 significant gonad-related genes were found, accounting for 3.59% of the total unique cDNAs. Specifically, the testis-specific LRR gene and testis-specific chromodomain Y-like protein gene were identified from fish for the first time. Six gonad-related microsatellite-containing ESTs were identified from the 129 ESTs containing 149 microsatellites. Expression patterns of 10 of these gonad-related gene homologues in ovaries and testes were examined by qRT-PCR. The results will be powerful resources for our further investigation to establish the molecular mechanisms of gonad development and sex differentiation in large yellow croaker.