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[1]苏 军** 管其龙 陈子强 陈在杰.水稻art1基因3’-UTR片段的克隆和验证*[J].应用与环境生物学报,2019,25(01):1-10.[doi:10.19675/j.cnki.1006-687x.2018.05023]
 SU Jun**,GUAN Qilong,CHEN Ziqian,et al.Cloning and validation of 3 ’-UTR fragments of rice art1 gene*[J].Chinese Journal of Applied & Environmental Biology,2019,25(01):1-10.[doi:10.19675/j.cnki.1006-687x.2018.05023]
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水稻art1基因3’-UTR片段的克隆和验证*()
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《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
25卷
期数:
2019年01期
页码:
1-10
栏目:
研究论文
出版日期:
2019-01-30

文章信息/Info

Title:
Cloning and validation of 3 ’-UTR fragments of rice art1 gene*
文章编号:
201805023
作者:
苏 军** 管其龙 陈子强 陈在杰
福建省农业科学院生物技术研究所,福建省农业遗传工程重点实验室 福州 350003
Author(s):
SU Jun** GUAN Qilong CHEN Ziqian CHEN Zaijie
Biotechnology Institute of Fujian Academy of Agricultural Sciences, Fujian Provincial Key Laboratory of Genetic Engineering for Agriculture, Fuzhou 350003, China
关键词:
水稻 3’-UTR 终止子 克隆 表达
Keywords:
rice 3-UTR terminator clone expression
DOI:
10.19675/j.cnki.1006-687x.2018.05023
摘要:
终止子是一段位于基因编码之后的序列,作为一种调控信号调控DNA的转录终止和RNA的释放,在基因工程技术应用中通常被构建在目的基因下游,用以调控基因的转录和表达。现有常用的终止子很少,克隆和验证新的终止子是植物基因工程技术发展的需要。本研究通过生物信息学分析和realtime-PCR表达验证,筛选出候选基因腺苷酸核糖基化作用因子(Similar to ADP-ribosylation factor1, arf1)基因,克隆了水稻art1基因3’-UTR。所克隆3’-UTR片段含有8个多聚信号元件和4个UE片段。将3’-UTR与gus基因融合后分别与玉米泛素启动子Ubiquitin和花椰菜花叶病毒( CaMV )启动子35S连接构建植物表达载体验证3’-UTR调控表达效果。烟草瞬时表达显示3’-UTR融合的gus 基因在35S和Ubi启动子驱动下,在烟草叶片中能正常表达;3’-UTR调控下的gus基因在水稻根、茎、叶、花和种子中均可稳定表达,且表达量与T-Nos终止子相当。研究表明:所克隆的3’-UTR可替代T-nos终止子,研究结果为植物基因工程技术的应用提供了新的调控元件。
Abstract:
Terminator is a sequence usually located at the end of a gene, as regulatory signals to block the transcription of DNA to RNA and release the transcript. At present, the commonly used terminators are very few, therefore cloning and validation of new terminators are necessary for the need of developing genetic modified crops. Here we clone a 3’-UTR sequence of candidate gene similar to ADP-ribosylation factor, arf1 and analyzed the control element of sequence. The sequence of arf1 3’-UTR contain 8 polyA sites and 4 U-rich elements. To confirm that the 3’-UTR of the arf1 gene is an important component for regulation of transcript stability, 2 reporter gene constructs were made by cloning the arf1 gene 3’-UTR downstream of the bacterial b-glucuronidase gene (uidA) under the control of the CaMV 35S and Ubi promoter respectively. Agrobacterium-mediated transient gene expression test show that 3’-UTR - fusion gus gene driven by 35S or Ubi promoter can normally expression in the tobacco leaves. The gus gene expression regulated by 3’-UTR was stable in rice roots, stems, leaves, flowers and seeds, and the expression level was comparable to that of T-Nos. The research shows that the clone 3’-UTR can replace T-Nos terminator, and the research results provide new regulatory elements for the application of plant genetic engineering technology.

备注/Memo

备注/Memo:
收稿日期: 2018-05-16 接受日期 Accepted: 2018-06-08
*福建省科技重大专项(2016NZ0001-2)和省属公益类科研院所科研专项(2018R1019-1)资助?
**通讯作者(E-mail: sj@fjage.org)
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更新日期/Last Update: 2018-07-13