原核增强子样序列的筛选及其应用研究  

Screening and application of prokaryotic enhancer-like sequence 3A

作　　者：韩峰[1] 何建新[1] 廖小慧[1] 王艳[1] 吴淑华[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所,北京100052

出　　处：《中华实验和临床病毒学杂志》2010年第3期175-177,共3页Chinese Journal of Experimental and Clinical Virology

基　　金：国家科委＂863＂高科技资助项目（102-11-03-03）

摘　　要：目的 从大肠埃希菌C600株染色体基因组中筛选原核增强子样序列,构建携带原核增强子样序列的表达载体,探讨其对干扰素基因表达的影响.方法 采用氯霉素乙酰转移酶基因（CAT）作为报告基因,从大肠埃希菌C600株染色体基因组中筛选具有原核增强子样活性的序列,构建携带增强子样序列的表达载体,表达干扰素基因和检测干扰素活性.结果 从大肠埃希菌C600株染色体基因组中筛选到一个原核增强子样序列3A,其正、反向增强活性分别能提高β-半乳糖苷酶活性7.11和2.93倍.证实它的增强活性体现在转录水平;用原核增强子样序列3A的功能区3P3构建的表达载体,其表达的IFN-α2b型干扰素比原表达载体活性高3.7倍.结论 从大肠埃希菌C600株染色体基因组中筛选到一个原核增强子样序列3A,携带有原核增强子样序列的表达载体可提高干扰素基因的表达水平.Objective To screen enhancer-like sequences from Escherichia coli strain c600 genome,to construct an expression vector harboring prokaryotic enhancer-like sequence and study the effect of interferon gene expression.Methods Enhancer-like element from Escherichia coli strain C600 genome was obtained by using the ehloramphenieol acetyl-transferase（CAT）gene as reporter gene.An expression vector harboring prokaryotic enhancer-like sequence from Escherichia coli strain C600 was constructed.Interferon was expressed and assayed.Results An enhancer-like sequences with distance and orientation independence property were screened and named 3A.Quantification test showed that the direct and reverse orientation of 3A could increase the activity of β-galactosidase with 7.11 and 2.93 times.The enhancing activity of the element was on transcription level.An expression vector harboring the prokaryotic enhancer-like sequence 3P3 which was enhancing function region of sequence 3A was constructed. Using this vector the antiviral activity of interferona-2b was increased by 3.7 times in comparison with the original expression plasmid.Conclusion 3A enhancer-like sequence was screened from Escherwhia coli strain C600 genome.Interferon gene was highly expressed by using an expression vector harboring enhancer-like sequences.

关 键 词：原核增强子样序列 表达载体 

分 类 号：R346[医药卫生—基础医学]