成花素基因PdFT的克隆及其对牡丹成花的影响

doi:10.3864/j.issn.0578-1752.2014.13.013

摘要/Abstract

摘要： 【目的】克隆紫牡丹FT同源基因，分析其表达模式以及对紫牡丹成花的调控作用。【方法】以紫牡丹为试验材料，通过RT-PCR的方法克隆紫牡丹FT同源基因PdFT。通过实时荧光定量PCR分析PdFT在紫牡丹不同组织、紫牡丹开花物候期各过程及不同处理的表达情况，并探讨PdFT与花芽发育状态的关系。同时，将克隆到的紫牡丹PdFT克隆到表达载体pET-28a上，构建融合表达载体pET-28a-PdFT，转化到大肠杆菌BL21（DE3）并诱导表达。【结果】克隆得到包括完整开放阅读框（ORF）的PdFT cDNA序列，ORF长度为522 bp，编码173个氨基酸，GenBank登录号为KF113360。氨基酸序列比对表明：PdFT蛋白具有一个典型的PEBP结构域，属于PEBP家族；并且与GenBank中已克隆的FT同源基因具有高度的同源性。实时荧光定量PCR分析结果表明，PdFT在紫牡丹根、茎、叶、芽中均有表达，芽中表达量最高，叶中表达量最低。紫牡丹开花物候期各过程PdFT的表达分析表明，在花芽膨大期表达量最高，随着花蕾的发育，PdFT的表达量逐渐降低。PdFT在不同光照温度条件下的表达分析表明，短日照和低温处理均抑制PdFT基因的表达。不同状态花蕾中PdFT的表达分析结果表明，败育花蕾PdFT的表达量低于正常花蕾。同时，GA3及去叶处理均能提高PdFT的表达量。所构建的原核表达载体，经IPTG诱导和SDS-PAGE电泳检测结果表明，表达蛋白与预期蛋白大小一致。【结论】紫牡丹PdFT与已克隆的FT同源基因高度同源，属于FT亚家族，在紫牡丹顶芽中表达量最高，可能调控牡丹开花，PdFT的克隆及表达分析为研究紫牡丹成花的分子机理奠定了基础。

关键词: 紫牡丹 , FLOWERING LOCUS T , 实时定量PCR , 原核表达

Abstract: 【Objective】The objective of this study was to clone the homolog of FT gene from tree peony and analyze the expression pattern and the potential function of flowering locus T (FT) of flowering in Paeonia delavayi Franch..【Method】The homologous FT gene, PdFT, was cloned by reverse transcript polymerase chain reaction (RT-PCR). The expression patterns of PdFT in different organs, developmental stages of flowering and treatments of P. delavayi Franch. were analyzed by using qRT-PCR and the relationship between the PdFT expression level and the different states of flower bud development was also studied. At the same time, PdFT was inserted into vector pET-28a and expressed in E. coli BL21 (DE3). 【Result】The full length of open reading frame (ORF) of PdFT was 522 bp, encoding 173 amino acid residues. The GenBank accession number is KF113360. Alignment of the AA sequences revealed that PdFT contains a conserved domain, which possesses the characteristics of the PEBP family. The result also suggested that PdFT shares high identity with FT proteins from other plants. qRT-PCR analysis of PdFT expression pattern in different organs showed that the highest expression levels were appeared in the buds of P. delavayi, followed by roots, stems and leaves. The expression pattern of PdFT from bud swelling to big-like flower-bud showed a decreased trend. The expression level of PdFT under long day condition was higher than that under short day condition. Cool treatment could decrease PdFT expression in roots and stems. The expression of PdFT under different floral conditions showed difference. The expression of PdFT in abortive flower buds was lower than that in normal bud. GA3 treatment and removing leaves could increase the expression of PdFT. The results of SDS-PAGE showed that the expressed protein in E. coli BL21 (DE3) was consistent with the size of expected. 【Conclusion】The PdFT isolated from P. delavayi Franch. is highly homologous with the other FT genes isolated from other species and belongs to FT subfamily. The higher expression of PdFT in buds suggested that it may play a role in regulating flowering in tree peony. PdFT gene cloning and expression analysis will lay a genetic basis for investigating the molecular mechanism of flowering in P. delavayi Franch.

Key words: Paeonia delavayi Franch. , FLOWERING LOCUS T , qRT-PCR , prokaryotic expression

朱富勇, 刘传娇, 薛璟祺, 王顺利, 张萍, 任秀霞, 张秀新. 成花素基因PdFT的克隆及其对牡丹成花的影响[J]. 中国农业科学, 2014, 47(13): 2613-2624.

ZHU Fu-Yong, LIU Chuan-Jiao, XUE Jing-Qi, WANG Shun-Li, ZHANG Ping, REN Xiu-Xia, ZHANG Xiu-Xin. Isolation of Florigen Gene PdFT and Its Effects on Flowering of Tree Peony（Paeonia delavayi Franch.）[J]. Scientia Agricultura Sinica, 2014, 47(13): 2613-2624.

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