应用基因表达系列分析(SAGE)技术研究高温处理前后家蚕的基因表达差异

Analysis on Expressional Difference of Silkworm Genes Before and After High Temperature Treatment by Using Serial Analysis of Gene Expression(SAGE)

高温对家蚕的生理和免疫能力有明显影响。为从分子水平上探究家蚕抗高温机制,应用基因表达系列分析(SAGE)技术获得家蚕5龄雌蚕高温处理前后中肠、丝腺和脂肪体组织的基因表达谱,构建高温组(34℃)和常温组(26℃)2个家蚕SAGE文库。2个家蚕SAGE文库分别包含3555107和3580976个原始标签,其中的标签种类分别为113684和131 296个,清洁标签的种类分别为45972和49467。比较2个文库清洁标签获得65 535种差异标签,共注释4249个基因,其中有1 062个差异表达的基因(P<0.05,错误检测率FDR≤0.001并且拷贝数的差异在2倍以上)。经GO分析发现2个文库中基因的分布极其相似,表明这些基因在不同的环境温度下有类似的生物学功能并参与类似的生理代谢过程。KEGG pathway分析显示有732个基因涉及176个KEGG路径,其中有40个为差异表达基因显著富集的路径(P<0.05),超过一半的路径与代谢、生物合成和信号传导有关。上述结果有助于对家蚕抗高温基因的鉴定以及探究基因调控的网络关系。

High temperature has obvious impact on physiology and immunization of silkworm. In order to explore the mechanism of silkworm resistance to high temperature at molecular level, serial analysis of gene expression （SAGE） technology was applied to investigate the expression profile of genes in midgut, silk gland and fat body of the 5th instar silkworm female larvae before and after high temperature treatment. Two silkworm SAGE libraries, one for 34 ℃ high temperature group and one for 26 ℃ normal temperature group, were constructed. The number of raw tags in the two silkworm SAGE libraries were 3 555 107 and 3 580 976 respectively, among which the number of tag types were 113 684 and 131 296, and the number of clean tags types were 45 972 and 49 467, respectively. A comparison on clean tags inthe two SAGE libraries revealed that there were 65 535 different tags. Totally 4 249 genes were annotated, and 1 062 of them had difference in expression level （P 〈 0.05, FDR〈0. 001 and multiple difference of copy number〉 2）. GO analysis showed that the distributions of genes in two libraries were highly similar, indicating that these genes have similar biological functions and are involved in similar physiological metabolic processes under different environmental temperatures. In addition, KEGG path- way analysis indicated that 732 genes were involved in 176 KEGG pathways, including 40 pathways rich in differentially expressed genes with significant difference （P 〈0. 05）. More than half of the pathways were related to metabolism, biosynthesis and signal transduction. The above results are conducive to the identification of high temperature resistance genes in silkworm and the investigation of gene regulation networks,