基因芯片技术对哮喘病人相关基因的筛选和分析

目的 利用基因芯片技术寻找哮喘病患者与正常人外周血单核细胞之间差异表达基因,拟为哮喘的早期诊断及预防提供分子标记.方法 用淋巴细胞分离液分别提取16例哮喘病患者与16例正常人外周血单核细胞,用QIAGEN Rneasy Kit提取纯化样本总RNA,并合成用荧光标记的cRNA,分别与含有41 000条基因序列的全基因芯片杂交,以基因表达倍数值≥2.0和基因表达倍数值≤-2.0为阈值来确定差异表达基因,然后用Genespring软件利用生物信息学方法对差异表达基因进行功能分类分析.结果 按P＜0.05差异显著性标准,从34 183条表达基因谱中,筛选出哮喘患者与正常对照差异表达2倍以上的基因有4177条,差异表达2倍以上已知与哮喘相关的基因有19条.经代谢途径分析发现这些差异基因主要涉及到炎症反应、免疫反应、防御反应、创伤反应、外部刺激反应等8大功能分类.结论 哮喘的发生涉及众多基因表达的改变,芯片技术可以有效地筛选出哮喘患者与正常对照的差异表达基因,对进一步探索哮喘的发病机制、有效的干预或逆转哮喘具有重要意义.

Screening and analysis of candidate gene expression in asthma using microarray technology

Objective To screen different gene expressions in peripheral blood mononuclear cells between asthma patients and normal people by DNA microarray in order to provide molecular markers for early diagnosis and prevention of asthma.Methods Participants with asthma(n=16) and healthy controls(n=16) underwent peripheral blood collection.Monocytes were isolated using Ficoll-Paque and extracted for total RNA by QIAGEN Rneasy Kit.Then we used the Cy3 to mark the cDNA of normal people and asthma patients respectively,and used the Agilent Homo microarray chips containing 41,000 genes/ESTs to screen gene expression differentially according to the screening criteria of Ratio≥2 and Ratio≤-2.At last,we used Gene spring software to analyze the biologic function of the candidate genes.Results By using this DNA microarray technology,4177 candidate genes in the asthma patients were found from 34183 target genes,which expressed twice higher than those in the normal people.19 gene expressions correlating to asthma were found with twice fold differences,as compared with the normal.Analysis showed that these differentially expressed genes mainly related to the following pathways like inflammation response,immune response,defense responses,wound responses and external stimuli.Conclusions Through screening and analysis of important candidate genes involved in asthma,we are able to investigate and explore the pathogenesis of asthma and to prevent asthma effectively in the future.