一个与小麦雄性不育育性转换相关的MADS-box转录因子基因

A MADS-Box Transcription Factor Related to Fertility Conversion in Male Sterile Wheat Lines

为了揭示YS型小麦温敏雄性不育育性转换的基础,构建了该类型不育系A3017的不育和可育幼穗正、反杂交的两个SSH-cDNA文库。经文库比较,在不育文库中筛选出一个与MADS-box基因同源的EST序列(GenBank登录号:36925702)。以该EST序列的同源性比对和拼接结果为依据,设计引物对该基因在可育和不育幼穗中的表达进行了RT-PCR分析,结果表明,该基因在不育幼穗中表达量较高,可育幼穗中表达量很低。对不育幼穗中扩增出的cDNA片段进行克隆测序,获得了666bp的cDNA序列。序列分析表明,该片段编码160个氨基酸,具有MADS-box转录因子的典型结构域K-box,被定名为TaMS-MADSbox,与一个小麦MADSbox转录因子基因WAG的氨基酸序列的相似性为94%。进一步以3种不同类型的小麦雄性不育系和保持系的幼穗cDNA为材料,利用半定量RT-PCR对该基因的表达模式分析发现也存在类似差异,该基因在不育系幼穗中表达量较高,而保持系幼穗中表达量较低。以上分析表明,该MADS-box转录因子基因的表达与小麦雄性不育系的育性转化相关,表达量高时表现雄性不育,表达量低时表现雄性可育。

Male sterility is one of the major characteristics to be used in heterosis utilization of crops, in which thermo-sensitive or photo-sensitive male sterility is very important for two-line hybridization due to the conversion of their male fertility under special weather conditions. YS type thermo-sensitive male sterile wheat （Triticum astivum L.） lines are applicable for heterosis use in the major wheat production areas of northern China. To investigate the molecular basis of male fertility conversion of YS type thermo-sensitive male sterile wheat lines, we constructed the sterile and fertile suppression subtractive hybridization （SSH） cDNA libraries respectively, using the cDNA of the male sterile or fertile young spikes from the same individual of one YS type thermo-sensitive male sterile wheat line A3017 under controlled male sterile or fertile conditions. Comparing the EST sequences between the two cDNA libraries, an EST （GenBank accession number： 36925702） highly similar to MADS box transcription factor gene was selected from the sterile SSH-cDNA library and used as the probe to search the dbEST. A pair of primers was designed based on the aligned sequence of highly homological EST sequences, and used to detect the expression difference of this gene between male sterile and fertile spikes via Reverse Transcriptase PCR （RT-PCR）. The results showed that the expression of this gene in male sterile spikes was much higher than that in fertile spikes. Then the RT-PCR fragment amplified from the male sterile spikes was cloned and sequenced, a cDNA sequence with 666 bp and encoding 160 amino acids was obtained. The cDNA fragment contained the typical K-box domain of MADS-box, and designated as TaMS-MADSbox. The deduced amino acids were 94% similar to WAG （BAC22939）, an MADS box transcription factor of wheat. The expression profiles of this MADS-box transcription factor gene in the male-sterile lines and their maintainers of three types of male sterile wheat lines were further analyzed via semi-quantitative RT-PCR. The result showed similar patterns that the expression of the MADS-box transcription factor gene in male-sterile lines was much higher than that in maintainers. It suggests that the expression of TaMS-MADSbox is related to the fertility conversion of male-sterile lines. The spikes are male-sterile under high-level expression of TaMS-MADSbox, while fertile under the low-level expression.