Isolation And Characterization Of The Albinism-related Genes (DEN-1,DEN-2) From Turbot Scophthalmus Maximus

Two gene fragments of DEN-1 (GenBank accession number DQ886390) and DEN-2 (GenBank accession number DQ886391), differentially expressed between the ocular skin tissues of normal and albino turbot, were isolated by mRNA differential display and sequenced. 22 DNA fragments related to albinism were isolated. The size of these fragments was 100-1200 bp in length. It was confirmed by RT-PCR method that the expression levels of both DEN-1 and DEN-2 were down - regulated in the ocular skin tissue of albino turbot. The sequence homology search of DEN-1 revealed high sequence homologies with the UGGT genes of zebrafish and cattle, and the Ugcgl1 gene of Norway rat. High sequence homologies of DEN-2 were observed with the eya4 gene (the eye absent homolog 4) from red jungle fowl, zebrafish, human, Norway rat, mouse and dog. The BLASTX results were consistent with the Blast results. The fragments showed no high degree of similarity to previously reported gene may be a novel gene. The further research on these sequence information are very useful to obtain the complete novel gene and to know the mechanism of albinism of turbot.Expression pattern analysis was performed by relative quantitative RT-PCR to analyze the expression levels of DEN-1 and DEN-2 in four kinds of skin tissues: ocular side of normal turbot (termed N1 skin), ocular side of albino turbot (termed A1 skin), blind side of normal turbot (termed N2 skin) and blind side of albino turbot (termed A2 skin). The optical densities of the signals of DEN-1 and DEN-2 normalized byβ-actin gene show that DEN-1 and DEN-2 have the similar expression pattern: N1>N2≈A2>A1. A 497 bp sequence which had high homologies with the eya4 gene of many animals were produced by the in silico cloning. Electronic expression profile analysis of DEN-1 and DEN-2 in the different tissues, organs and development stages will provide valuable references for the further studies.