鸭Ccna1启动子克隆、序列分析及在鸭骨骼肌中的发育性表达  

Molecular Cloning,Bioinformatics Analysis of the Duck Ccna1 Gene Promoter Region,and Its Differential Expression Profiles in Skeletal Muscle Tissues

作　　者：杨振[1] 李彦莹[1] 罗俊[1] 张涛[1] 刘贺贺[1] 蒲梅[1] 肖香君 任艳霖 

机构地区：[1]四川农业大学动物遗传育种研究所,温江611130

出　　处：《畜牧兽医学报》2017年第2期377-384,共8页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金(31301964);大学生创新性实验项目(1510626040)

摘　　要：为研究Ccna1基因在鸭骨骼肌发育中的作用,本研究应用RT-PCR扩增和克隆了鸭Ccna1基因启动子区并作生物信息学分析,应用荧光定量PCR技术检测Ccna1基因以及参与该基因表达调控的多个肌源性转录因子在鸭胚骨骼肌组织发育过程中的表达模式。结果表明,扩增得到鸭Ccna1基因启动子2 213bp。序列分析表明,鸭Ccna1启动子存在典型的TATA-box、CAAT-box调控元件,有MyoD、MRF4、MyoG、Sp1、PITX1及MEF2等多个转录因子结合位点。定量结果发现,MyoD、MRF4、MyoG和Ccna1在鸭胚胎骨骼肌发育过程中均有表达。在鸭胸肌中,MRF4在E23时期表达量最高,显著高于D8的表达量(P<0.05);在腿肌中,MRF4和MyoG表达量在E17时期达到最高,且显著高于D2、D8(P<0.05)。聚类分析表明,在鸭胸肌组织中Ccna1表达模式与MyoD一致,在腿肌组织中Ccna1表达模式与MRF4、MyoG一致。初步推测,Ccna1参与到了鸭骨骼肌组织的发育,且在鸭腿肌的生长发育中,MRF4、MyoG可能与Ccna1相互作用进而调控骨骼肌发育,而在鸭胸肌组织中,Ccna1的表达可能与MyoD的转录调控有关。To investigate the roles of Ccna1 gene in duck skeletal muscle development,its promoter sequence was amplified by RT-PCR,then was analyzed using bioinformatics methods.qRTPCR was used to detect the expression profiles of Ccna1 during duck embryonic muscle development,as well as the expression profiles of myogenic transcription factors predicted to take part in regulation of Ccna1 transcription.We obtained 2 213 bp promoter region of duck Ccna1 gene.Bioinformatics analysis showed there were typical TATA-box,CAAT-box and binding sites for transcription factors MyoD,MRF4,MyoG,Sp1,PITX1 and MEF2in the Ccna1 gene promoter region.The qRT-PCR data showed that MyoD,MRF4,MyoGand Ccna1 expressed in duck muscle tissues at embryonic development stages.In the breast muscle,the expression of MRF4 was the highest in E23 and was significantly higher than that in D8（P0.05）.In the leg muscle,the expression of MRF4 and MyoG were the highest in E17,and was significantly higher than that inD2,D8（P0.05）.Clustering results showed that the expression patterns of Ccna1 was consistent with MyoDin breast muscle,while in the leg muscle,Ccna1 was clustered together with MRF4 and MyoG.It was preliminary concluded that Ccna1 was involved in the duck skeletal muscle development,and Ccna1 might be associated with the MRF4 and MyoG during the leg muscle development,whereas in the breast muscle development,its expression might be regulated by MyoD.

关 键 词：鸭 Ccna1 启动子序列 生物信息学分析 聚类 骨骼肌 

分 类 号：S834[农业科学—畜牧学] S813.1[农业科学—畜牧兽医]