基于RNA-seq技术开发中间偃麦草基因组特异分子标记

中间偃麦草(Thinopyrum intermedium,2n=6x=42,JJJSJSStSt或Ee Ee EbEbStSt)是小麦遗传改良的重要近缘植物,高抗锈病、白粉病等病害,耐逆性强。开发中间偃麦草基因组特异分子标记对于加快其优异基因向普通小麦中的转移和利用具有重要意义。为建立快速准确开发中间偃麦草基因组特异分子标记的新体系,首先利用RNA-seq技术对中间偃麦草及其基因组供体二倍体长穗偃麦草、百萨偃麦草、假鹅观草的苗期叶片进行转录组测序,然后根据测序结果设计EST-SSR引物,对普通小麦、中间偃麦草及其基因组供体材料进行DNA扩增分析,从中筛选鉴定中间偃麦草基因组特异标记。结果表明,在所合成的200对ESTSSR引物中,有75对引物(37.5%)在中间偃麦草、二倍体长穗偃麦草、百萨偃麦草和假鹅观草中具有特异扩增,引物多态率较高。说明利用RNA-seq技术开发EST-SSR引物可以高效地用于筛选中间偃麦草基因组特异分子标记,这一技术体系也可应用于其他作物及其近缘物种的特异分子标记开发。

Thinopyrum intermedium; Wheat; RNA-seq; Specific molecular markers

Thinopyrum intermedium,one of the important relatives of common wheat,has good tolerance to environmental stress and resistance to diseases such as rust and powdery mildew.Development of specific molecular markers is valuable for transformation and application of the favorable genes from Th.intermedium.In order to develop an efficient method of screening the specific molecular markers in Th.intermedium,the RNA-seq data of Th.intermedium and its genome donors,including Th.elongatum,Th.bessarabicum and Pseudoroegneria strigosa were used for developing EST-SSR markers.Two hundred pairs of EST-SSR primers were synthesized and verified through amplifying different DNA materials,and 75(37.5%) primers have specific amplification in Th.intermedium,Th.elongatum,Th.bessarabicum and Ps.strigosa with good polymorphism.The result indicates that developing EST-SSR primers from RNA-seq would be efficient in screening specific molecular markers of Th.intermedium,which can be used in developing specific molecular markers for other crops and their related species.