抗纹枯病、赤霉病的转TaPIEP1基因小麦的分子鉴定与选育

摘要

小麦纹枯病（主要病原为禾谷丝核菌）和赤霉病（主要病原为禾谷镰刀菌）已成为我国小麦生产的重要病害.TaPIEP1是从小麦中分离到的1个病原诱导的基因,其编码蛋白是可与GCC-box顺式元件结合、转录激活型的ERF转录因子.本研究以8个转TaPIEP1基因小麦株系的T4和T5代植株为试材,进行了外源转TaPIEP1基因的PCR检测、Southern杂交、RT-PCR与Q-RT-PCR的分析以及纹枯病菌、赤霉病菌接种与抗性鉴定.结果表明,外源TaP IEP1基因在转基因小麦中能够稳定遗传,以单拷贝或双拷贝整合到7个转基因小麦株系基因组的不同位点；外源TaPIEP1基因在转基因小麦中能超量表达；与受体扬麦12相比,TaPIEP1表达水平高的8个转基因小麦株系对纹枯病抗性显著提高,4个株系中一些材料对赤霉病抗性显著提高,3个株系中一些材料兼抗纹枯病和赤霉病,说明TaPIEP1正向参与了小麦对纹枯病和赤霉病抗性反应,利用该基因通过基因工程可创制抗纹枯病、赤霉病的小麦新种质.%Wheat sharp eyespot, mainly caused by Rhizoctonia cerealis, and Fusarium head blight (FHB), primarily caused by Fusarium graminearum, are important diseases of wheat (Triticum aestivum L.) in China. We have isolated a pathogen-induced ERF gene from wheat, TaPIEP1, which encodes the ERF transcription factor TaPIEP1. TaPIEP1 localizes to the nucleus, binds to the GCC-box cis-element and possesses the transcriptional-activation activity. To study the roles of TaPIEP1 in wheat defense responses to the major pathogens of sharp eyespot and FHB, we characterized the TaPIEP1 transgenic wheat plants in T4 and T5 generations by PCR, Southern blot, RT-PCR, and Q-RT-PCR analyses. We also evaluated the disease resistance in these TaPIEP1 transgenic plants through inoculating R. Cerealis and F. Graminearum. The PCR and Southern blotting results showed that the alien TaPIEP1 was inherited stably in transgenic wheat plants, and was independently integrated with a single copy or two copies into seven transgenic wheat lines, suggesting that these transgenic wheat lines derived from seven transformants. The RT-PCR and Q-RT-PCR analysis results indicated that the alien gene TaPIEP1 was over-expressed in eight transgenic wheat lines. Compared with untransformed wheat host Yangmai 12, the eight transgenic wheat lines over-expressing TaPIEP1 showed significantly-enhanced resistance to R. Cerealis infection. Out of them, some plants of three transgenic wheat lines displayed improved-resistance to both R. Cerealis and F. Graminearum infections. These results suggest that TaPIEP1 gene is involved in defense responses to attack with R. Cerealis and F. Graminearum, and TaPIEP1 is useful for improving wheat resistance to both dis-eases.