利用抑制消减杂交技术(SSH)研究与Ty21a免疫相关的基因

Isolation of genes that are induced by Ty21a in intestinal cells by suppression subtractive hybridization (SSH)

目的:利用抑制消减杂交技术（SSH）筛选与Ty21a免疫相关的新基因。方法:以Ty21a免疫小鼠的肠细胞为tester,以正常小鼠的肠细胞为diver,提取mRNA,反转录构建cDNA文库,利用SSH技术筛选Ty21a免疫相关的新基因。结果:通过SSH技术和cDNA微矩阵技术,发现了7条与GenBank中已公布的表达序列标签（expressed sequence tag,EST）片段没有明显同源性,可能为Ty21a免疫相关的新基因。其中3条正在用RACE-PCR法进行获得全长cDNA的工作。结论:SSH技术是一种高效的差异基因筛选方法,基因表达谱芯片技术是高通量进行基因表达模式研究的方法,Ty21a可诱导多种免疫相关新基因的表达。

Objective:To isolate the genes induced by Ty21a. Methods: Prepare the intestinal cells of Balb/C mouse with and without Ty21a immunization, extract the polyA+ RNA, synthesize the dscDNA by using reverse transcription-PCR,get the differentially expressed genes by suppression subtractive hybridization(SSH).Results:Seven genes were novel genes revealed by GenBank searching whose expression were probably induced by Ty21a were isolated by SSH and cDNA microarray. The full length cDNA sequence of three genes are now obtaining through RACE-PCR. Conclusion: SSH is an effective method for isolating the differentiatlly expressed genes; High throughput cDNA microanay is a credibility method to profile changes in gene expression; Ty21a can induce the expression of some new genes related to immune.