Borrás-Hidalgo Orlando, Thomma Bart P H J, Collazo Cyrelys, Chacón Osmany, Borroto Carlos J, Ayra Camilo, Portieles l Roxana, López Yunior, Pujol Merardo
Laboratory of Plant Functional Genomics, Center for Genetic Engineering and Biotechnology, P.O. Box 6162, Havana, 10600, Cuba.
Mol Plant Microbe Interact. 2006 Apr;19(4):399-406. doi: 10.1094/MPMI-19-0399.
In order to identify tobacco (Nicotiana megalosiphon) genes involved in broad-spectrum resistance to tobacco blue mold (Peronospora hyoscyami f. sp. tabacina), suppression subtractive hybridization was used to generate cDNA from transcripts that are differentially expressed during an incompatible interaction. After differential screening by membrane-based hybridization, clones corresponding to 182 differentially expressed genes were selected, sequenced, and analyzed. The cDNA collection comprised a broad repertoire of genes associated with various processes. Northern blot analysis of a subset of these genes confirmed the differential expression patterns between the compatible and incompatible interaction. Subsequent virus-induced gene silencing (VIGS) of four genes that were found to be differentially induced was pursued. While VIGS of a lipid transfer protein gene or a glutamate decarboxylase gene in Nicotiana megalosiphon did not affect blue mold resistance, silencing of an EIL2 transcription factor gene and a glutathione synthetase gene was found to compromise the resistance of Nicotiana megalosiphon to P. hyoscyami f. sp. tabacina. Potentially, these genes can be used to engineer resistance in blue mold-susceptible tobacco cultivars.
为了鉴定参与对烟草霜霉病（烟草霜霉病菌）具有广谱抗性的烟草（大虹吸烟草）基因，采用抑制性消减杂交技术从在不相容互作过程中差异表达的转录本中生成cDNA。通过基于膜的杂交进行差异筛选后，选择了与182个差异表达基因对应的克隆，进行测序和分析。该cDNA文库包含了与各种过程相关的大量基因。对这些基因的一个子集进行Northern印迹分析，证实了亲和互作和不相容互作之间的差异表达模式。随后对发现差异诱导的四个基因进行了病毒诱导的基因沉默（VIGS）。虽然大虹吸烟草中脂质转移蛋白基因或谷氨酸脱羧酶基因的VIGS不影响对霜霉病的抗性，但发现EIL2转录因子基因和谷胱甘肽合成酶基因的沉默会损害大虹吸烟草对烟草霜霉病菌的抗性。这些基因有可能用于培育对霜霉病敏感的烟草品种的抗性。
