Division of Plant Microbe Interactions, CSIR-National Botanical Research Institute, Rana Pratap Marg, Lucknow, 226001, India.
Mol Biol Rep. 2014 Mar;41(3):1385-99. doi: 10.1007/s11033-013-2983-7. Epub 2014 Jan 10.
The inoculation of Pseudomonas putida NBRIC19 protected wheat plant from phytotoxic effect of Parthenium hysterophorus (Parthenium) and enhanced root length, shoot length, dry weight, spike length and chlorophyll content. With the aim to screen for genes differentially expressed in P. putida NBRIC19-inoculated wheat grown along with Parthenium (WPT), the suppression subtractive hybridization (SSH) methodology was employed. The SSH analysis was performed with WPC (uninoculated wheat grown along with Parthenium) as driver and WPT as tester. The cDNA library, enriched with differentially expressed ESTs (expressed sequence tags), were constructed from WPT. Following an initial screen of 165 ESTs in our library, 32 ESTs were identified, annotated and further validated by semiquantitative RT-PCR. The differentially expressed ESTs were associated with general stress response, defense response, growth and development, metabolic process, photosynthesis, signal transduction, and some other with unknown function. Five ESTs showing downregulation in expression level in response to Parthenium got upregulated due to P. putida NBRIC19 inoculation and further validated by quantitative real time PCR analysis at different time intervals viz. 15, 30, 45 and 90 days. SSH has been implemented for the first time to gain insights into molecular events underlying successful role of P. putida NBRIC19 in providing protection to wheat against Parthenium. The information generated in this study provides new clues to aid the understanding of genes corresponding to differentially expressed ESTs putatively involved in allelopathic interactions. Further characterization and functional analysis of these genes may provide valuable information for future studies of the molecular mechanism by which plants adapt to allelopathic effect of Parthenium.
铜绿假单胞菌 NBRIC19 接种可保护小麦植株免受豚草（Parthenium）的植物毒性影响，并增加根长、茎长、干重、穗长和叶绿素含量。为了筛选铜绿假单胞菌 NBRIC19 接种与豚草（WPT）一起生长的小麦中差异表达的基因，采用抑制性消减杂交（SSH）方法。SSH 分析以 WPC（未接种的与豚草一起生长的小麦）为驱动，WPT 为测试。从 WPT 构建了富含差异表达 EST（表达序列标签）的 cDNA 文库。在我们的文库中对 165 个 EST 进行初步筛选后，鉴定了 32 个 EST，并通过半定量 RT-PCR 进一步验证。差异表达的 ESTs 与一般应激反应、防御反应、生长发育、代谢过程、光合作用、信号转导有关，还有一些与未知功能有关。由于铜绿假单胞菌 NBRIC19 的接种，5 个 ESTs 的表达水平下调，对豚草的反应上调，并通过不同时间点（15、30、45 和 90 天）的定量实时 PCR 分析进一步验证。这是首次实施 SSH，以深入了解铜绿假单胞菌 NBRIC19 在为小麦提供对豚草的保护方面发挥作用的分子事件。本研究提供的信息为辅助理解差异表达 EST 所涉及的基因提供了新线索，这些基因可能涉及化感相互作用。这些基因的进一步表征和功能分析可能为未来研究植物适应豚草化感作用的分子机制提供有价值的信息。
