摘要：

从甜荞中分离出FaesANT基因,研究其在同型长花柱(LH)甜荞中的表达模式及功能,揭示其在同型长花柱甜荞花器官发育调控中的作用,为甜荞分子育种与种质创新提供基因资源。结合PacBio SMRT(单分子实时测序)转录测序和RACE技术从甜荞中克隆FaesANT基因全长,石蜡切片结合qRT-PCR技术分析FaesANT基因在同型长花柱甜荞中表达的组织特异性和花芽形态分化过程中表达量的动态变化;并结合同型长花柱甜荞VIGS后花器官的表型解析基因的功能。结果表明:甜荞FaesANT基因全长1 982 bp,包含1 632 bp ORF,编码543个氨基酸,分子系统发育树分析将其聚为真双子叶植物ANT进化系。FaesANT在雄蕊和雌蕊以及幼果中表达量较高,但在三者中的表达量差异不显著,在茎、叶和花被片中仅能检测到微弱转录信号;在花芽分化过程中,FaesANT基因在雌雄蕊原基分化后有明显的表达,并在雄蕊花丝伸长期表达量达到峰值,随着花芽的进一步发育和分化,FaesANT的表达量逐渐下降,至开花时降至最低。在TRV2-FaesANT处理的强表型同型长花柱植株的花表现出雌蕊花柱和雄蕊花丝缩短、雄蕊数量减少,部分雄蕊花药花瓣化,FaesANT基因在这些强表型植株花中的表达量极显著降低。结合FaesANT表达模式和基因沉默后花器官表型分析,推测FaesANT主要参与甜荞花丝和花柱长短调控,同时也参与了甜荞雄蕊数量的调控。

关键词: 甜荞, ANT同源基因, 基因表达, 基因沉默, 同型长花柱

Abstract:

Research on the expression pattern and function of FaesANT gene in long-homostyle(LH)common buckwheat and uncovering its roles involved in floral organ development will help to provide gene resources for the molecular breeding and germplasm innovation of buckwheat.The full-length FaesANT gene was isolated from long-homostyle common buckwheat by combining PacBio SMRT(Single Molecule Real-Time)sequencing and RACE technique.FaesANT expression levels in different tissues and floral buds at sequential developmental stages of long-homostyle common buckwheat were separately detected by using paraffin sections and qRT-PCR.Moreover,the FaesANT gene function involved in floral development was investigated by using virus-induced gene silencing(VIGS)in long-homostyle common buckwheat.The results were as follows,FaesANT cDNA(1 982 bp)comprised a 1 632 bp ORF encoding a 543 amino acid transcription factor.Phylogenetic tree analysis grouped the FaesANT into the eudicots ANT lineage.FaesANT expression levels were mainly detected in stamen,pistil and young fruit of long-homostyle common buckwheat and showed no significant differences among them.However,only weak transcripts were detected in the stems,leaves and tepals.During floral bud differentiation,obvious expression of FaesANT gene was detected when pistil and stamen primordia formation,and FaesANT expression level reached peak during filament elongation stage of stamen.Then,FaesANT expression decreased gradually with the floral bud development and dropped to the lowest level after flowering.TRV2-FaesANT-treated long-homostyle plants with strong phenotypic changes produced flowers with style-and filament-length decreased,and stamen number reduced.In addition,some TRV2-FaesANT-treated long-homostyle plants displayed part stamens with anther homeotically converted into tepaloid structure.Moreover,FaesANT expression decreased significantly in the flowers of FaesANT-silenced long-homostyle plants with strong phenotypic changes.It suggested that FaesANT was mainly involved in regulating filament and style length,and determining stamen number of common buckwheat based on the gene expression pattern and phenotypes of VIGS-treated long-homostyle common buckwheat flowers.

Key words: Fagopyrum esculentum, ANT homologous gene, Gene expression, Gene silence, Long homostyle

中图分类号: 

• S517荞麦