Friedrich Miescher Institute, PO Box 2543, CH-4002 Basel, Switzerland.
Mol Plant Pathol. 2001 May 1;2(3):125-34. doi: 10.1046/j.1364-3703.2001.00059.x.
Summary Suppression Subtractive Hybridization (SSH) was applied in a search for genes induced during the compatible interaction between Phytophthora infestans and potato. Using potato leaves that had been treated with benzo(1,2,3)thiadiazole-7-carbothioic acid S-methylester (BTH) as the control tissue, a low redundancy library with a relatively low frequency of the classic plant Pathogenesis-Related (PR) genes was generated. 288 of the clones were screened for induced sequences using Inverse Northern analysis (hybridizing the arrayed clones with radiolabelled cDNA populations). Of the 75 clones that were detectable by this method, 43 appeared to be induced. Eleven of these clones were then analysed by total RNA blot analysis, and elevation of transcript levels during P. infestans infection was confirmed for 10 of them. Some of the cDNAs analysed by RNA blot analysis have homology to genes already known to be induced during infection, e.g. to beta-1,3-glucanase. Another group of cDNAs have homology to enzymes involved in detoxification: gamma-glutamylcysteine synthetase, cytochrome P450, glutathione S-transferase and an MRP-type ABC transporter. Other infection induced cDNAs encode putative proteins that have not previously been reported to be induced by infection: e.g. the ER-located chaperone BiP, and a homologue of Aspergillus nidulans SudD, which was isolated as a suppressor of a mutation in chromosome disjunction. The differential library therefore presents the opportunity to analyse the metabolic changes occurring during infection, and the disease process itself in more detail.
抑制差减杂交（SSH）被应用于寻找在马铃薯晚疫病菌与马铃薯的亲和互作过程中诱导产生的基因。利用用苯并（1,2,3）噻二唑-7-羧酸 S-甲酯（BTH）处理过的马铃薯叶片作为对照组织，构建了一个低冗余文库，其中经典的植物病程相关（PR）基因的出现频率相对较低。通过反向 Northern 分析（用放射性标记的 cDNA 群体杂交排列的克隆）筛选了 288 个克隆，以寻找诱导序列。通过这种方法可检测到的 75 个克隆中，有 43 个似乎被诱导。然后用总 RNA 印迹分析对其中的 11 个克隆进行分析，证实其中的 10 个在晚疫病菌感染过程中转录水平升高。用 RNA 印迹分析分析的一些 cDNA 与已知在感染过程中被诱导的基因具有同源性，例如β-1,3-葡聚糖酶。另一组 cDNA 与参与解毒的酶具有同源性：γ-谷氨酰半胱氨酸合成酶、细胞色素 P450、谷胱甘肽 S-转移酶和 MRP 型 ABC 转运体。其他感染诱导的 cDNA 编码以前未报道过被感染诱导的假定蛋白：例如内质网定位的伴侣蛋白 BiP，以及 Aspergillus nidulans SudD 的同源物，该基因作为染色体分离突变的抑制因子被分离出来。差异文库因此提供了分析感染过程中发生的代谢变化以及更详细的疾病过程的机会。
