黄瓜cDNA芯片的构建及其在黄瓜缺镁胁迫下基因差异表达研究中的应用

Construction of a Cucumber cDNA Microarray and Its Application in the Study of Response of Cucumber Plants to Magnesium Deficiency Stress

基因芯片技术在研究植物基因功能中发挥着越来越重要的作用,本试验通过GenBank搜索已发布的生物代谢途径中的关键酶基因序列,设计特异性引物,采用RT-PCR法扩增这些酶的相应基因片段,在完成432个基因片段的克隆分离、测序和生物信息学分析工作的基础上研制出国内首张黄瓜cDNA芯片。该芯片含有9个质控cDNA片段和423个cDNA探针,涉及光反应、卡尔文循环、碳水化合物代谢、水-水循环、信号传导、激素代谢、光呼吸、防御、蛋白与氨基酸代谢等多个代谢途径。利用该芯片对黄瓜缺镁胁迫下基因表达谱进行了初步研究,并发现在缺镁胁迫下差异表达的22个基因,其中10个基因的表达受缺镁处理抑制,12个基因的表达受缺镁处理诱导。该芯片的研制为进一步研究黄瓜功能基因的高通量时空变化提供了有效的技术支撑。

Microarray technology plays increasing roles in the study of plant gene function. By using BLAST biosoftware and data from GenBank, the specific primers for key metabolism enzyme genes in cucumber were designed, and the specific fragments amplified by RT-PCR were cloned and sequenced. A total of 432 cDNA sequences corresponding to genes involved in photochewistry, carbon fixation, carbohydrate metabolism, water-water cycle, signal transduction , hormone metabolism, defense, photorespiration and amino acid metabolism were obtained and categorized. At last, a cDNA microarray with 423 cDNA fragments and 9 quality control cDNA fragments were built. The microarray was then used to study the changes of gene expression profiles of cucumber plants after exposure to magnesium starvation. The results showed that 10 genes were down regulated, while 12 genes were up regulated in magnesium deficiency plants. This microarray will provide us a powerful tool for the study of gene function in response to a series of stresses.