东亚三角涡虫DjWnt5a克隆及在胚胎不同发育阶段中的表达  

Cloning and Expression of the DjWnt5a Gene in Planarian Dugesia Japonica during Different Embryonic Development Stages

作　　者：甄辉[1] 吴素格[1] 鹿晴晴[1] 赵博生[1] 

机构地区：[1]山东理工大学生命科学学院发育与进化生物学实验室,山东淄博255049

出　　处：《中国生物化学与分子生物学报》2016年第2期199-206,共8页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金项目(No.31572263);山东省自然科学基金项目(No.ZR2013CM011)~~

摘　　要：WNT(wingless-type MMTV integration site family)蛋白是一类分泌性糖蛋白家族,Wnt5a为非经典Wnt蛋白的典型代表,在各种器官的发育中发挥重要作用。为了探索东亚三角涡虫Dj Wnt5a基因在涡虫胚胎不同发育阶段中的表达和功能,利用RT-PCR(reverse transcription PCR)和RACE(rapid-amplification of c DNA ends)技术克隆并分析了Dj Wnt5a c DNA的表达。结果表明,Dj Wnt5a基因由1 523 bp组成,编码371个氨基酸,其中含有23个保守的半胱氨酸结构域。利用MAGE 5.2构建进化树分析发现,东亚三角涡虫与地中海涡虫的Wnt5a聚群,并位于两侧对称动物的基部,可能是Wnt5a的原始类型。利用整体原位杂交技术分析Dj Wnt5a基因的表达,结果显示,Dj Wnt5a基因在涡虫发育的第三阶段开始表达,主要集中在胚胎咽及胚带;随着胚胎发育直至完全孵化出幼虫,Dj Wnt5a基因逐步在神经原基和中枢神经系统表达。由此推断,东亚三角涡虫Dj Wnt5a基因在涡虫胚胎发育时,参与神经原基的形成和分化,是涡虫胚胎神经发育必不可少的基因之一。Wnt proteins are a family of highly conserved secreted glycoproteins that regulate cell-to-cell interactions during embryogenesis. Wnt5 a is a typical representative of non-canonical Wnt protein,which plays an important role in the development of various organs. To explore the expression and function of the Dj Wnt5 a during the embryonic development in planarian Dugesia japonica,the Dj Wnt5 a c DNA was cloned by RACE and the expression pattern was detected by Whole-mount in situ hybridization. The results showed that the Dj Wnt5 a was comprised of 1 523 bp and was translated in one open-reading frame to give a protein of 371 amino acids. Constructing phylogenetic tree using MAGE5. 2 analysis revealed that planarians Dj Wnt5 a protein falling at the base of bilatera,suggesting that planarians Dj Wnt5 a was the archetype of the bilatera. Expression analysis of Dj Wnt5 a gene indicated that the Dj Wnt5 a m RNA was first found in stage 3 of the embryonic development,expressing in the embryonic pharynx and germ band.Along with the embryonic development,Dj Wnt5a-positive embryonic cells were increased,and were distributed in the neural primordiums and the central nervous system of the fully hatched larvae. These results suggested that Dj Wnt5 a is one of the essential genes in the embryonic nervous development,playsa vital role in the formation and differentiation of the neural primordium during the planarians embryonic development.

关 键 词：东亚三角涡虫 DjWnt5a基因 整体原位杂交技术 胚胎发育 

分 类 号：Q52[生物学—生物化学] Q71