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目的研究铁皮石斛(Dendrobium officinale)种子接菌共生萌发差异基因表达谱特征。方法采用抑制差减杂交(SSH)技术,分别以接菌及未接菌培养5周的铁皮石斛种子cDNA作为检测子与驱赶子,构建铁皮石斛种子接菌共生萌发抑制性消减cDNA文库。结果对部分克隆产物测序,经GenBank中BLASTx同源比较后,进行基因功能注释,得到100个具有植物同源性的表达序列标签(EST),主要涉及细胞与染色体结构、RNA合成、信号转导、能量与代谢、蛋白质合成与降解及细胞防御等。随机挑选5个目标基因,实时荧光定量PCR分析发现这5个基因在共生萌发的种子中均上调表达。结论铁皮石斛种子接菌共生萌发涉及多种途径相关基因表达调控,本实验为兰科植物种子萌发分子机制研究奠定基础。
Objective To study the differential gene expression profiles of Dendrobium officinale seed germination. Methods Suppression subtractive hybridization (SSH) technique was used to detect the cDNA library of symbiotic germination of Dendrobium candidum by using the cDNA of D. officinale as inoculation and non-inoculation for 5 weeks respectively. Results Some of the cloned products were sequenced. After homologous comparison with BLASTx in GenBank, gene function annotation was performed and 100 ESTs were obtained, which mainly involved in cell and chromosome structure, RNA synthesis, signal transduction , Energy and metabolism, protein synthesis and degradation and cell defense. Five target genes were randomly selected and analyzed by real-time fluorescence quantitative PCR. All the five genes were upregulated in the symbiotic germination seeds. Conclusion The symbiotic germination of Dendrobium candidum seedlings involved in the regulation of gene expression in many ways. This experiment laid the foundation for the molecular mechanism of seed germination of Orchidaceae.