基因芯片技术鉴定棉铃虫胁迫后棉花差异表达基因

摘要

In general, gene expression changes when cotton is infested by phytophagous insects. Screening and identification of these regulatory genes induced by cotton bollworm (Helicoverpa armigera Hiibner) infestation will help us understand the molecular mechanism of the host plants resistance to insect pests. In this paper, cotton {Gossypium spp.cv. Zhongl2) was as test material. The cotton plants were divided into control groups and pest stress treatment groups (including 6, 12, 24 and 48 h, pest stress treatments, respectively). The total RNA were extracted from control groups and different pest stress treatments, and the Affymetrix cotton gene chip analysis were applied to explore gene expression profiles. It was found that a total of 4 109 genes (28%) in leaves weredifferentially expressed after induced 6 h, 1 917 of them was up-regulated and 2 192 of them was down-regulated, respectively. After 12 h, a total of 2 605 genes(l 8%) were differentially expressed, 1 326 of them was up-regulated and 1 279 of them was down-regulated, separately. After 24 h, a total of 3 213 genes (22%) were differentially expressed, 1 424 of them is up-regulated and 1 789 of them was down-regulated, respectively. After 48 h, a total of 2 763 genes (19%) were differentially expressed, 1 450 of them was up-regulated and 1 313 of them is down-regulated, separately. Bioinformatics analysis revealed that function of some differentially expressed genes involved roxidative stress response, defense response, signal transduction, transcriptional regulation, flavone and flavonol biosynthesis, terpenoids synthesis and metabolism, biosynthesis of alkaloids derived from many kinds of amino acids, etc. Furthermore, some potential genes which regulate specific volatiles emission in cotton were also identified. It was found that the gene expression of farnesyl pyrophosphate synthase was regulated by circadian rhythm. Meanwhile, (+)-delta-cadinenesynthase gene expression level increased when cotton plants was infested by H. Armigera. Real-time quantitative PCR results indicated that farnesyl pyrophosphate synthase gene and chalcone-flavonone isomerase gene had the highest expression level when cotton infested 6 h by H. Armigera. However, farnesyl pyrophosphate synthase gene and chalcone-flavonone isomerase gene had the lowest expression level when cotton infested 48 h by H. Armigera. These findings proide base studying data for defense mechanism when cotton is infested by cotton bollworm, and help us further understand the molecular regulation of terpenoids synthesis under phytophagous insects stress.%棉花在受到植食性昆虫为害后基因表达会发生变化,筛选签定这些调控基因有助于解析昆虫诱导棉花抗虫性的分子机制.本研究以棉花(Gossypium spp.)(中12)为实验材料,分别从正常生长条件下的棉花(对照)和棉铃虫(Helicoverpa armigera Hübner)取食胁迫6、12、24和48 h的棉花叶片(处理)中提取总RNA,采用Affymetrix棉花基因芯片进行了基因表达谱分析.结果表明,棉铃虫取食6h时,差异表达基因共有4 109个(占28％),其中上调1 917个,下调2 192个；棉铃虫取食12h时,差异表达基因共有2 605个(占18％),其中上调1 326个,下调1 279个；棉铃虫取食24 h时,差异表达基因共有3 213个(占22％),其中上调1 424个,下调1 789个；棉铃虫取食48 h时,差异表达基因共有2 763个(占19％),其中上调1 450个,下调1 313个.进行生物信息学分析后发现,这些基因功能涉及氧化应激响应、防御响应、信号转导、转录调控、黄酮类生物合成、萜类化合物合成与代谢以及源于多种氨基酸的罗勒生物碱生物合成等多个方面.另外,从芯片结果中鉴定获得调控棉花特异性挥发物的相关基因,发现法呢基焦磷酸合酶基因的表达量受昼夜节律的影响,同时,棉铃虫取食棉花诱导杜松萜烯合酶基因表达量上调.实时荧光定量PCR检测发现,棉铃虫取食6h的棉株中法呢基焦磷酸合酶基因和查尔酮异构酶基因表达量较高,而在被棉铃虫取食48 h的棉株中这两个基因的表达量较低.研究结果为棉花受棉铃虫为害后产生的防御机制的研究提供了基础资料,进一步为植食性昆虫取食诱导植物产生萜烯类化合物提供了依据.