合浦珠母贝Pf-Cnot2基因的克隆与表达分布

采用RACE技术从合浦珠母贝外套膜组织中克隆得到了CCR4-NOT复合体第二亚基(Cnot2)基因,命名为PfCnot2。该基因全长2 616 bp,开放阅读框为1 725 bp,编码574个氨基酸,理论分子量为61 ku,理论等电点为6.25。蛋白序列分析表明它富含丝氨酸、甘氨酸和苏氨酸,具有保守的NOT2_3_5结构域。系统进化分析显示Pf-Cnot2蛋白与脊椎动物的Cnot2蛋白进化关系较远。Pf-Cnot2在合浦珠母贝的外套膜边缘、外套膜缘膜、足、生殖腺、内脏团、闭壳肌和鳃等组织中均有表达,在生殖腺中表达量最高。该基因从担轮幼虫期至幼贝期均有表达,担轮幼虫期表达量最少,D型期比担轮幼虫期有显著增高。上述研究结果为进一步了解Pf-Cnot2在合浦珠母贝中发挥的作用提供了基础数据。

Cloning and expression of Pf-Cnot2 in Pinctada fucata

Cnot2, a subunit of CCR4-NOT, was cloned from the mantle tissue of Pinctada fucata by RACE PCR and named Pf-Cnot2. The complete sequence was 2616 bp long and the open reading frame was 1725 bp, which encoded 574 amino acid residues with the theoretical molecular weight of 61 ku and theoretical isoelectric point of 6.25. Protein sequence analysis showed that it was rich in serine, glycine and threonine, and had a conservative Not2_3_5 domain. Phylogenetic analysis indicated that Pf-Cnot2 had a far evolutionary relationship with Cnot2 of vertebrate. This gene was expressed in tissues of mantle edge, mantle pallial, foot, gonad, visceral mass, adductor muscle and gill in Pinctada fucata, in which, gonad had the highest expression level, followed by gill and mantle pallial. Pf-Cnot2 was expressed during the whole time since trochophore developed into the juvenile shell, and got to its lowest level at trochophore stage. D-shape stage was significantly higher than trochophore stage. The results above provided basic material for further study of the role that Pf-Cnot2 played in Pinctada fucata.