利用cDNA-AFLP分析纹枯病菌诱导的玉米差异表达基因

cDNA-AFLP analysis of Rhizoctonia solani AG1-IA-induced maize differential gene expression

以纹枯病菌AG1-IA(Rhizoctonia solani)诱导玉米高耐纹枯病自交系R15,采用cDNA-AFLP技术分析其基因差异表达谱。在拔节期对R15幼苗进行接菌处理,12、24、36、48、60 h分别取材,以不接菌为对照。用56对AFLP选扩增引物对处理和对照的cDNA进行AFLP分析,得到87个差异片段,回收并剔除假阳性,克隆获得18条阳性差异条带(TDFs)。BLASTn比对结果表明,其中可以找到同源序列的有13个TDFs,按其功能可分为信号传导(2个)、抗病与防御基因(2个)、转录调控(2个)、能量代谢(2个)等。对13个TDFs基因进行了半定量RT-PCR分析,结果表明13个差异片段在对照与处理,或是处理的不同时段存在着表达量的差异。

cDNA amplified fragment length polymorphisms（cDNA-AFLP） was used to analyze gene expression profile in high tolerance maize inbreed lines R15 induced by AGI-IA （Rhizoctonia solani）. At the join- ring stage, the seedlings were inoculated and sampled at 12, 24, 36, 48 and 60 h and the untreated seedlings were U＇god as control. Gene expression profile was tested by cDNA-AFLP. From the obtained 87 differential fragments, eliminating the false positive, 18 transcript-derived fragments （TDFs） were cloned. The sequences were then analyzed through bioinformatical methods, 13 of them had significant homology sequence with that in GenBank database and their function were as follows： signal transduction （two sequences）, resistance and defense （ two sequences）, transcription and regulation （ two sequences）, energy metabolism （ two sequences） etc. 13 differential expression fragments were confirmed by semi-quanfitive RT-PCR at different inoculating times.