Molecular Cloning and Expression Analysis of Short Stature Homeobox 2 (Shox2) in Oryctolagus cuniculus
CHEN Rui;ZHU Zifeng;HUANG Jian;YANG Xiaonong;College of Life Science and Technology,Southwest Minzu University;Key Laboratory of Ministry of Education and Sichuan Province for Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization;Chengdu Huitai Biomedicine CO.;Hospital of Southwest Minzu University;
To clone and characterize short stature homeobox 2 (Shox2)gene in New Zealand rabbit,predict its structure and function,and to investigate its expression pattern,the full-length CDS sequence of Shox2 was cloned,and the biological characteristics of Shox2 protein were analyzed in this study,such as homology,hydrophilicity,secondary and tertiary structure characteristics and phylogenetic tree.Western blot was employed to verify fusion protein TrxA-Shox2 .Quantitative real-time RT-PCR(RT-qPCR)was employed to detect the expression patterns of Shox2 in kidney,aorta,heart,dermis,lung,brain,liver,spleen,gallbladder and muscle tissues at 4different developmental phases(E20.5d,E28.5d,10dand adult)of 3animals for each phase.Shox2 gene was successfully cloned in present study(GenBank accession number:KP726285),CDS of Shox2 was composed of 666bp encoding 221amino acids.According to the results of bioinformatics analysis,Shox2 belonged to the unstable alkaline protein,61.99%of which amino acids was alpha helix,33.48%for random coil,4.52%for extended strand.The result of Western blot verified the expression of fusion protein TrxA-Shox2 .The analysis of homology and phylogenetic tree showed that the amino acid sequence of Oryctolagus cuniculus Shox2 protein had high homology and close genetic distance with tarsier,gorilla,rattus norvegicus and human,which meant that Shox2 gene was conservative during evolutionary process.According to the result of RT-qPCR,Shox2 gene universally expressed in most tissues detected in this study.A relatively higher Shox2 mRNA expression level was detected in kidney,gallbladder and muscle tissues in embryo for E20.5d,in heart and spleen in embryo for E28.5d,in dermis and liver for 10d,and in derminds and spleen for adult.In this study,Shox2 gene was successfully cloned and characterized.Shox2 gene universally expressed in all tissues detected,its expression level was related to the function of Shox2 in different tissues and at different developmental stages,which was strictly regulated.
【Fund】： 国家重大支撑项目课题(2014BAD13B03);; 西南民族大学研究生学位点建设项目(2014XWD-S0906)
【CateGory Index】： S829.1
【CateGory Index】： S829.1