高羊茅光周期调控基因FaCONSTANS亚细胞定位与差异表达分析

Subcellular Localization and Differential Expression Analysis of FaCONSTANS Gene in Tall Fescue

构建高羊茅光周期调控基因FaCONSTANS与绿色荧光蛋白基因GFP融合的植物表达载体p-FaCONSTANS-hGFP,基因枪转化法转入洋葱表皮细胞,荧光显微镜检测融合基因的瞬时表达。并运用实时荧光定量PCR研究光周期调控基因FaCONSTANS在长日照与短日照条件下昼夜24 h的表达差异。结果表明:FaCONSTANS基因编码的蛋白产物位于细胞核,符合它作为转录因子特性;在长日照和短日照条件下,光周期调控基因FaCONSTANS在1个光周期内表达均是由低到高再降低,长日照条件下FaCONSTANS基因的转录峰在有光照时出现,短日照条件下FaCONSTANS基因的转录峰在黑暗中出现,但短日照处理条件下的转录峰比长日照处理条件下转录峰早2 h出现,约高2.3倍。由此说明,FaCONSTANS基因的表达受光周期调控,与生物钟控制的昼夜节律有关。

The plant expression vectors p-FaCONSTANS-hGFP with a green fluorescent protein gene has been constructed and transformed into onion skin cells by gene gun method,transient expression of fusion gene was detected with fluorescence microscopy.We detected FaCONSTANS gene transcription levels in tall fescue in day and light under long and short days condition by real-time fluorescence quantitative PCR.The result showed that FaCONSTANS protein was located in the nucleus which is in line with the characteristics of transcription factors.A expression peak was detected in both long days and short days condition.Transcription peak of FaCONSTANS gene occurs in light under long-days condition,transcription peak of FaCONSTANS gene occurs in the dark under short-day condition.However,the level of FaCONSTANS transcription peak was significantly higher（2.3 fold） under short days than that under long days,the peak of FaCONSTANS transcription level under short days condition is earlier 2 hours than that of long days condition.This indicates that FaCONSTANS gene expressed with a circadian rhythm and is regulated by photoperiod.