锦鲤酪氨酸酶基因序列分析及其在不同锦鲤品系不同组织中的表达

Analysis of tyrosinase gene and tissue expression in five different strains of Koi carp (Cyprinus carpio Koi)

为了解色素控制基因与体色分布的关系,实验用分子克隆技术得到了锦鲤酪氨酸酶(tyrosinase,Tyr)cDNA基因序列,并利用半定量PCR的方法分析了酪氨酸酶基因在锦鲤不同品系和组织中的表达差异。实验得到了长约1 779 bp锦鲤酪氨酸酶cDNA基因,其包括长1 608 bp开放阅读框,编码535个氨基酸。cDNA、蛋白水平序列分析和系统发育分析都表明酪氨酸酶在鱼类间的保守性要高于鱼类与其他脊椎动物间的保守性。半定量分析研究显示,酪氨酸酶基因在皮肤、肝、心、脑和眼中都有表达,其中眼部和黑色皮肤表达最高,其次是脑,红色和黄色皮肤,肝和心的表达最低。不同品系同组织间眼、肝、心、脑的酪氨酸酶基因表达差异不显著,但是在皮肤中有差异表达,黑色皮肤表达最高,其次是红色和黄色皮肤,在白色皮肤中也有少量表达。酪氨酸酶基因在非黑色组织中有较高表达可能与存在多种形式酪氨酸酶有关,也有可能非黑色素细胞能转化为黑色素细胞。

Black is an important body color of Koi （Cyprinus carpio Koi） which is performed through melanin synthesized by melanocyte, and tyrosinase is an important rate-limiting enzyme in melanin synthesis. It＇s unclear of genetic mechanism of Koi body color. The present work obtained Koi tyrosinase （Tyr） gene sequence and investigated the expression of Tyr gene in different strains and tissues using semi-quantitative PCR, trying to reveal the relationship between the pigment control genes and the body color. The results showed 1 779 bp of Tyr gene, including a 1 608 bp open reading frame encoding 535 amino acids. Sequence analysis of eDNA and protein and phylogenetic analysis indicated that conservation of Tyr is higher in teleost than between teleost and other vertebrates. Semi-quantitative analysis showed the highest Tyr expression in the eye and black skin, intermediate in brain, red and yellow skin, and the lowest in liver and heart. There was no significant difference of Tyr expression between the same tissues of eyes, liver, heart, brain from different Koi carps, but differentiated in the skins with the highest in black skin, followed by red and yellow skin and small amount in white skin. The different expression was related to tissue function. The role of tyrosinase in non-melanoma cells was unclear. There possibly exist multiple forms of tyrosinase, some of which is inhibited, or the xanthophores and erythrophores could be translated into functional melanoeytes.