板栗CmMYB转录因子基因的克隆及在花序发育中的表达分析

CmMYB cloning and its expression during catkin differitiation of castanea mollissima

板栗大量雄花序影响产量形成,寡雄性状是板栗重要的育种目标。利用RT-PCR与RACE扩增方法,从板栗极短雄花序芽变和正常雄花序cDNA中分离出一个板栗MYB转录因子cDNA全长,进行测序和序列分析。结果表明,克隆的cDNA片段总长为1 522 bp,基因内部含有完整的开放阅读框架,大小为1 071 bp,可编码长度为357个氨基酸残基的蛋白质,所推导的蛋白质氨基酸序列与苹果的MYB91、豌豆的MYB转录因子和蒺藜苜蓿的MYB转录因子基因分别具有88%、87%和87%的氨基酸序列同源性。序列分析及空间结构预测发现该序列具有完整的R2R3区域,推测其具有转录激活功能。命名为CmMYB(GenBank登录号为GU076490)。实时荧光定量PCR结果显示,芽变雄花序CmMYB转录因子在雄花序原基形成期、花簇原基形成期、花朵原基形成期、雄蕊原基形成期和花药形成期的表达量均低于正常雄花序中的表达量,而在花被原基形成期表达量高于正常雄花序中表达量,说明该转录因子调控的主要基因与雄花序的短化存在一定联系。

Chestnut yield has been limited due to a great mass of male flowers. One important aim in chestnut breeding is to select the cultivar with less male flowers and high ratio of burs. In the experiment, the full-length cDNA encoding MYB transcription factor was isolated from the normal and mutant short catkin of Castanea mollissima and amplified by reverse transcription polymerse chain reaction （RT-PCR） and rapid-amplificatlon of eDNA ends （RACE）. The results showed that MYB was I 522 bp, containing a 1 071 bp open reading frame （ORF） which encoded a protein of 357 amino acids with 88%, 87% and 79% similarity in amino acid sequence conmpared with that encoded by MYB in Maltts domestica, Pisum sativum and Medicago truncatula, respectively. The gene was named CmMYB （accession numbers GUO76490）. Sequence analysis and spatial structure prediction revealed that the sequence had a complete R2R3 area and speculated that transcription activation. Realtime RT-PCR indicated that： in the primordia formation phase of catkin, floral cluster, flower, androeci- um and anther, the expression of CmMYB gene in mutant catkins was lower than that in the normal catkins. However, at the stage of perianth primordium formation （programmed cell death happened at the same time）, the expression of CmMYB gene in mutant catkins was much higher than that in the normal. It was speculated that the gene regulated by CmMYB was relative to shorting of mutant catkins.