Tbx5基因在漠斑牙鲆（Paralichthys olivaceus）变态前的表达

鲽形目鱼类仔鱼经过变态后形成眼睛和偶鳍等器官左右不对称的稚鱼。Tbx家族成员参与脊椎动物心脏、肢体和眼等器官的发育，而Tbx基因与鲽形目鱼类变态的关系目前尚无报道。我们运用扫描电镜和软硬骨染色等方法对出膜后2d（变态早期）和26d（变态前期）的牙鲆仔鱼的形态进行观察，牙鲆出膜后2d胸鳍原基出现，出膜后26d胸鳍发育成熟，而在出膜后26d胸鳍和腹鳍仍呈现左右对称。我们克隆了牙鲆Tbx5基因的CDS序列，该序列编码的多肽具有部分T-box保守结构域，与斑马鱼Tbx5同源性高达93%。系统进化分析显示：这段CDS序列与硬骨鱼的Tbx5基因聚为一支，但与四足动物Tbx5相分离。整体原位杂交显示出膜后26d的牙鲆仔鱼中，Tbx5基因不仅在胸鳍、眼背侧、脊柱和尿囊表达，且在胸鳍和眼背侧的表达呈现左右不对称，左侧面（有眼侧）的表达量均高于右侧面（无眼侧）的表达量。综上结果暗示，牙鲆Tbx5基因可能参与了变态过程中眼睛与胸鳍左右不对称的形成。研究亮点： 鲽形目鱼类都经历变态发育过程，产生显著的左右不对称形态，但调控该过程的分子机制尚不清楚。本文对牙鲆变态初期仔鱼进行了形态学观察，并对牙鲆Tbx5进行克隆及其表达分析，并探讨Tbx5与牙鲆仔鱼眼睛与胸鳍不对称变态过程的关系，为进一步研究牙鲆变态机制奠定基础。

The expression of Tbx5 gene at pro-metamorphosis stage in southern flounder, Paralichthy solivaceus

Flatfish larvae metamorphose from bilaterally symmetrical larvae into asymmetrically shaped juveniles with two eyes in ocular side, and the ocular side pectoral fin is larger than the one on blind side. Many members of Tbx family take part in many organogenesis processes, including heart, limb and eye. However, the relationship between Tbx gene and flatfish larvae metamorphosis has not been reported. We used scanning electron microscopy (SEM) and bone/cartilage staining method to observe the morphology in Paralichthys olivaceus pre-metamorphosis stage (2 dph) and pro-metamorphosis stage (26 dph) larvae. The results indicate that pectoral fins bud appears at 2 dph, and pectoral and pelvic fins mature at 26 dph. The pectoral and pelvic fins are still bilaterally symmetric by that stage. Then we cloned partial CDS of Tbx5 gene, which encoded a polypeptide with conserved T-domain, sharing 93% identity with zebrafish Tbx5. Phylogentic analysis reveals that this partial CDS is grouped with teleost branch, but separated with tetrapod branch. The whole-mount in situ hybridization shows that Tbx5 gene is expressed in pectoral fins, eyes, vertebra, and allantoic vesicle at 26 dph larvae of Paralichthys olivaceus. In addition, the expression of Tbx5 gene in eyes and pectoral fins are asymmetry between the left and the right sides, which is higher expressed in the eyes and pectoral fins of the left side (ocular side) than the right side (blind side). All these results suggest that Tbx5 genes may involve in the metamorphosis process of Paralichthys olivaceus.