僧帽牡蛎EGFR基因的克隆及早期发育阶段的表达

Cloning and Expression of EGFR in Embryos and Larvae of Crassostrea angulata

僧帽牡蛎(Crassostrea angulata)也称葡萄牙牡蛎,是我国广泛分布的一种传统大宗养殖贝类.本研究利用消减杂交文库结合RACE技术,从僧帽牡蛎中克隆获得了表皮生长因子受体(Epidermal growth factor receptor,EGFR)的cD-NA全长序列.EGFR基因cDNA序列1 608 bp,编码298个氨基酸,预测存在1个跨膜结构域和5个酪蛋白激酶II磷酸化位点.构建了该基因的UPGMA进化树,比对发现其氨基酸序列与高等动物的差异较大.给出了该基因的标准曲线,利用RT-PCR技术检测了该基因在僧帽牡蛎胚胎发育早期阶段的表达量变化.该基因从僧帽牡蛎发育的卵到D型幼虫期间均有表达.其中早期的卵母细胞和后期的担轮幼虫,D型幼虫阶段表达量较高,表明该基因在牡蛎幼体胚胎发育早期具有阶段性调控的作用.

Crassostrea angulata was a shellfwash with widely dwastribution in China,and was a traditional aquaculture species of high yield.In this study,the authors cloned epidermal growth factor receptor（EGFR） cDNA full sequences of C.angulata by RACE technology combining with oyster SSH library.The sequence was 1 608 bp,encoding 298 amino acids,which had a transmembrane domain and five casein kinase II phosphorylation sites.By BLAST analysis,the authors constructed the UPGMA cladogram of amino acid sequence of EGFR from different species.The amino acid sequence quite different with higher animals such as human.The authors constructed the EGFR′s standard curve,and used Real-time PCR to detect the gene changes in the 8 early stages of embryonic development of C.angulata.The 8 stages are egg,zygote,4 cell,morula,blastula,gastrula,trochophore,and D-veliger.The EGFR had expression from the egg to the D-veligar stages.In the egg and trochophore,D-veligar stage there was a relatively high expression of EGFR mRNA.The results indicated that EGFR had a stage regulation mode in the embryos and larvae of C.angulata.In early development time the embryos relied on mRNA in the egg.After gastrula stage,the EGFR expression increased,might playing an important role in the cleavege and organ conformation.