苦荞黄酮转运相关蛋白基因FtAH4L的克隆及表达分析

为研究苦荞黄酮转运相关基因,以苦荞(Fagopyrum tataricum)品种"西荞二号"为材料,克隆到1条质膜H+-ATPase基因(autoinhibited H+-ATPase isoform 4 like,AH4L),将其命名为FtAH4L。通过开放阅读框(ORF)分析,FtAH4L基因cDNA全长3 398bp,开放阅读框2 898bp,编码966个氨基酸残基,理论分子量为109kD,等电点6.48。氨基酸保守基序比对分析表明,AH4L在植物种间较为保守。在茉莉素诱导处理和5种光(白色荧光、LED白光、LED蓝光、LED红光和UV-B)处理芽期苦荞后,采用半定量RT-PCR和AlCl3比色法分析结果表明,茉莉素处理后的苦荞胚轴和子叶中FtAH4L基因表达量与黄酮含量均显著上升,且二者呈正相关关系;5种光对子叶中FtAH4L表达量无显著影响,但均显著增加其黄酮含量;胚轴中,除LED红光外,各种光均显著提高FtAH4L表达量和总黄酮含量,且LED蓝光与UV-B的影响极显著。该研究结果为深入研究FtAH4L基因参与苦荞黄酮转运奠定了基础。

Cloning and Expression Analysis of Flavonoids Transporter Gene FtAH4L from Fagopyrum tataricum

The gene FtAH4L,which encoding the autoinhibited H⁺-ATPase isoform 4 like(AH4L) involved into flavonoids transportation,was cloned from tartary buckwheat cultivar Xiqiao No.2(Fagopyrum tatarium).The length of FtAH4L gene was 3 398 bp,and the ORF(Open Reading Frame) was 2 898 bp encoding 966 amino acids.The molecular weight of FtAH4L was 109 kD,and its pI was 6.48.Conserved motifs and phylogenetic analysis demonstrated that AH4Ls are relatively conservative in different plant species.At the seedling stage of tartary buckwheat,semi-quantitative RT-PCR analyzed the expression profiles of FtAH4L gene,and AlCl₃ colorimetric method measured the total flavonoids content in cotyledons and hypocotyls,after jasmonates treatment and different lighting(White fluorescent,LED White,LED Blue,LED Red and UV-B).The results showed that FtAH4L expression and flavonoids content not only had a significant increase(P<0.05),but also showed a positive correlation in cotyledons and hypocotyls after jasmonates treatment.Under all the lightings,FtAH4L expression was unremarkable change(P>0.05) while flavonoids content was raised obviously in cotyledons.However,in hypocotyls,FtAH4L expression and flavonoids content were promoted notably(P<0.05) except LED Red.Particularly,LED blue and UV-B presented the extremely significant affect(P<0.01).This work could provide an important reference to understand the mechanism of FtAH4L gene involved in flavonoids transportation in tartary buckwheat.