花生生長素響應(yīng)因子基因AhARF3的克隆與表達(dá)分析

唐桂英　崔維佩　徐平麗　李鵬祥　朱潔瓊　單雷

摘要：生長素響應(yīng)因子（auxin response factor， ARF）是調(diào)控生長素響應(yīng)基因表達(dá)和信號轉(zhuǎn)導(dǎo)的一類轉(zhuǎn)錄因子，在植物生長發(fā)育的各個階段均起重要的調(diào)節(jié)作用。本研究基于花生種子萌發(fā)前后轉(zhuǎn)錄表達(dá)差異分析，克隆到1個生長素響應(yīng)因子基因AhARF3。序列分析結(jié)果顯示，該基因的ORF區(qū)為2 115 bp，編碼704個氨基酸，推測相對分子量為72.92 kD，等電點(diǎn)6.86。不同器官表達(dá)譜分析結(jié)果顯示，AhARF3為組成性表達(dá)，莖中的表達(dá)量最高，根與花中的表達(dá)量較低且近似，干種子中最低。種子萌發(fā)過程的表達(dá)分析顯示，萌發(fā)10～72 h 的種子中AhARF3表達(dá)量均比干種子中極顯著增加，尤其是萌發(fā)48～72 h，AhARF3的表達(dá)水平不僅比萌發(fā)前期各階段種子中的大幅上調(diào)，與根、莖、葉、花及各發(fā)育階段的種子相比，也具有明顯的表達(dá)優(yōu)勢。

關(guān)鍵詞：花生;生長素響應(yīng)因子;基因表達(dá);萌發(fā)

中圖分類號：S565.2：Q785文獻(xiàn)標(biāo)識號：A文章編號：1001-4942（2019）09-0050-06

Cloning and Expression Analysis of AhARF3 Gene in Peanut

Tang GuiyingCui Weipei2， Xu PingliLi Pengxiang2， Zhu Jieqiong2， Shan Lei2

（1. Biotechnology Research Center， Shandong Academy of Agricultural Sciences/Shandong Provincial Key

Laboratory of Crop Genetic Improvement， Ecology and Physiology， Jinan 250100， China; 2. College of Life Sciences，

Shandong Normal University， Jinan 250014， China）

Abstract Auxin response factor （ARF） is a kind of important transcription factor in regulating the expression of auxin response genes and signal transduction， which is involved in various development processes in plant. In this study， the cDNA of AhARF3 gene was cloned and analyzed based on the comparative analysis of transcriptomic data between dry seeds and newly germinated seeds. Sequence analysis results showed that the ORF length of AhARF3 was 2 115 bp， which encoded 704 amino acids， and the relative molecular weight was estimated to be 72.92 kD and the isoelectric point was 6.86. The results of qRT-PCR showed that AhARF3 was expressed by the constitutive type， the expression level was the highest in stems and the lowest in dry seeds. The expression level of AhARF3 in seeds germinated for 10～72 hours was significantly higher than that in dry seeds， especially 48～72 hours after germination， and was not only significantly higher than that in pre-germination stage， but also had obvious expression advantages compared with roots， stems， leaves， flowers and seeds at the other developmental stages.

Keywords Peanut （Arachis hypogaea L.）; Auxin response factor （ARF）; Gene expression; Germination

生長素響應(yīng)因子（auxin response factor， ARF）是一類能夠調(diào)節(jié)生長素響應(yīng)基因表達(dá)的轉(zhuǎn)錄因子，它通過特異性識別并結(jié)合生長素應(yīng)答元件，激活或抑制基因的表達(dá)[1]。ARF蛋白一般包含3個結(jié)構(gòu)域：N端的B3類DNA結(jié)合域（DNA binding domain， DBD）、中間的MR結(jié)構(gòu)域-抑制結(jié)構(gòu)域（repression domain， RD）或激活結(jié)構(gòu)域（activation domain， AD）和C端的二聚化作用的結(jié)構(gòu)域（carboxy-terminal dimerization domain， CTD）[2]。ARF的功能與植物細(xì)胞內(nèi)生長素的濃度密切相關(guān)，當(dāng)生長素濃度較低時，生長素響應(yīng)基因Aux/IAA與ARF形成異源二聚體，ARFs的作用被抑制;當(dāng)生長素濃度升高到一定水平，Aux/IAA經(jīng)泛素化途徑快速降解，ARFs被釋放并與生長素響應(yīng)元件TCGTCTC結(jié)合，生長素響應(yīng)基因被激活或抑制，植物體隨之發(fā)生一系列生長素反應(yīng)[3]。……