College of Life Sciences, Northwest A&F University, Yangling, Shaanxi, 712100, PR China.
BMC Genomics. 2009 Dec 8;10:586. doi: 10.1186/1471-2164-10-586.
Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat worldwide. To establish compatibility with the host, Pst forms special infection structures to invade the plant with minimal damage to host cells. Although compatible interaction between wheat and Pst has been studied using various approaches, research on molecular mechanisms of the interaction is limited. The aim of this study was to develop an EST database of wheat infected by Pst in order to determine transcription profiles of genes involved in compatible wheat-Pst interaction.
Total RNA, extracted from susceptible infected wheat leaves harvested at 3, 5 and 8 days post inoculation (dpi), was used to create a cDNA library, from which 5,793 ESTs with high quality were obtained and clustered into 583 contigs and 2,160 singletons to give a set of 2,743 unisequences (GenBank accessions: GR302385 to GR305127). The BLASTx program was used to search for homologous genes of the unisequences in the GenBank non-redundant protein database. Of the 2,743 unisequences, 52.8% (the largest category) were highly homologous to plant genes; 16.3% to fungal genes and 30% of no-hit. The functional classification of all ESTs was established based on the database entry giving the best E-value using the Bevan's classification categories. About 50% of the ESTs were significantly homologous to genes encoding proteins with known functions; 20% were similar to genes encoding proteins with unknown functions and 30% did not have significant homology to any sequence in the database. The quantitative real-time PCR (qRT-PCR) analysis determined the transcription profiles and their involvement in the wheat-Pst interaction for seven of the gene.
The cDNA library is useful for identifying the functional genes involved in the wheat-Pst compatible interaction, and established a new database for studying Pst pathogenesis genes and wheat defense genes. The transcription patterns of seven genes were confirmed by the qRT-PCR assay to be differentially expressed in wheat-Pst compatible and incompatible interaction.
由小麦条锈菌（Puccinia striiformis f. sp. tritici，Pst）引起的小麦条锈病是全球范围内小麦最具破坏性的病害之一。为了与宿主建立亲和性，Pst 形成特殊的侵染结构，以最小化对宿主细胞的损伤侵入植物。尽管已经使用各种方法研究了小麦与 Pst 之间的亲和互作，但对互作分子机制的研究仍然有限。本研究旨在建立一个受 Pst 感染的小麦 EST 数据库，以确定参与亲和性小麦-Pst 互作的基因转录谱。
从接种后 3、5 和 8 天收获的感病小麦叶片中提取总 RNA，用于构建 cDNA 文库，从中获得了 5793 条高质量的 EST，聚类为 583 个 contigs 和 2160 个 singletons，得到了一组 2743 个 unisequences（GenBank 登录号：GR302385-GR305127）。BLASTx 程序用于在 GenBank 非冗余蛋白质数据库中搜索 unisequences 的同源基因。在 2743 个 unisequences 中，52.8%（最大类别）与植物基因高度同源；16.3%与真菌基因同源，30%无命中。所有 ESTs 的功能分类是基于数据库条目使用 Bevan 的分类类别建立的，该条目给出了最佳 E 值。大约 50%的 EST 与编码具有已知功能的蛋白质的基因显著同源；20%与编码具有未知功能的蛋白质的基因相似，30%与数据库中的任何序列没有显著同源性。定量实时 PCR（qRT-PCR）分析确定了 7 个基因的转录谱及其在小麦-Pst 亲和互作中的作用。
cDNA 文库可用于鉴定参与小麦-Pst 亲和互作的功能基因，并为研究 Pst 致病基因和小麦防御基因建立了新的数据库。通过 qRT-PCR 试验证实了 7 个基因的转录模式在小麦-Pst 亲和和不亲和互作中存在差异表达。
