摘要
目的分析肿瘤远处转移相关蛋白Ezrin mRNA的一、二级结构,寻找其各种反义技术作用的靶点。方法利用RNAdraw和RNAstructure两种计算机程序对Ezrin mRNA进行分析计算,得到其一、二级结构信息,选择两种程序计算出的共同的连续4个以上碱基未配对的单链成环区作为反义技术的靶区域,设计反义寡核苷酸(AsODNs)、核酶(Ribozyme)、脱氧核酶(DNAzyme or Deoxyribozyme)和小干扰RNA(SiRNA)等4种常用反义技术的作用靶点,再通过计算机程序OligoWalk利用最低自由能原则进行筛选,最后得到各反义技术的作用靶点。结果各反义技术的靶点数目分别为:AsODNs17个,Ribozyme5个,DNAzyme15个,SiR-NA10个;其中AsODNs靶点中较为理想的为G1749-A1771和C1777-G1794,Ribozyme为A2359-G2360,DNAzyme为A1789-U1790、A1756-G1757及A1784-G1785,SiRNA为G802-G808、G1749-A1771及A2354-G2360,并且A2354-G2360是所有4种反义技术共同的作用区域,可以作为一个通用靶点。结论正确的靶点设计为应用反义技术抑制Ezrin表达从而控制恶性肿瘤的远处转移提供了有效的指导作用。
【Objective】 Ezrin has been proved to play a significant role in the metastasis of malignant tumors.Antisense molecules can be inaccessible when the complementary target sequences of mRNA are sequestered in secondary structure.So the antisense technologies are often ineffective.To identify the primary and secondary structure and effective target sites of Ezrin mRNA for antisense technology is the purpose of this study.【Methods】 Two RNA folding computer programs,RNAdraw and RNAstructure,were used to predict the Ezrin mRNA primary and secondary structure.Single strand regions of at least four consecutive unpaired bases in Ezrin mRNA secondary structure were chosen to be the target sequences.The target sites which the four classes antisense molecules including antisense oligonucleotides（AsODNs）,Ribozyme,DNAzyme and short interfering RNA（SiRNA） binding to were designed according to these single strand regions.These designed target sites were screened by program OligoWalk which based on the principle of minimum free energy.The low minimum free energy of antisense molecule-target sequence duplexes was selected as the result.【Results】 AsODNs had 17 target sites,Ribozyme had 5 target sites,DNAzyme had 15 target sites,SiRNA had 10 target sites.Among these target sites,G1749-A1771 and C1777-G1794 were the perfect target sites of AsODNs because of the lower free energy of AsODNs-target sites duplexes.In the same way,A2359-G2360 was the perfect target site of Ribozyme,A1789-U1790,A1756-G1757 and A1784-G1785 were the perfect target sites of DNAzyme,G802-G808,G1749-A1771 and A2354-G2360 were the perfect target sites of SiRNA.Furthermore,A2354-G2360 was the common target site of the four antisense technologies.【Conclusion】 These results may provide a rational,efficient method for Ezrin mRNA knockdown and inhibit the metastasis of malignant tumors with antisense technology.
出处
《中国现代医学杂志》
CAS
CSCD
北大核心
2011年第4期377-383,388,共8页
China Journal of Modern Medicine
基金
高等学校博士学科点专项科研基金(No:20060558018)
关键词
EZRIN
二级结构
反义技术
靶点
ezrin
secondary structure
antisense technology
target site
