细叶百合LpAGD14基因的克隆与表达分析

doi:10.11686/cyxb2017317

草业学报 ›› 2018, Vol. 27 ›› Issue (2): 117-123.DOI: 10.11686/cyxb2017317

细叶百合LpAGD14基因的克隆与表达分析

汪王, 田忠平, 苏小霞, 杨柳慧, 周蕴薇^*

东北林业大学园林学院,黑龙江哈尔滨150000

收稿日期:2017-07-24 修回日期:2017-10-18 出版日期:2018-02-20 发布日期:2018-02-20
通讯作者: dlzhyw@126.com
作者简介:汪王(1993-),女,安徽安庆人,硕士。E-mail:1192763410@qq.com
基金资助:
国家自然科学基金项目(31470698)资助

Cloning and expression analysis of LpAGD14 from Lilium pumilum

WANG Wang, TIAN Zhong-ping, SU Xiao-xia, YANG Liu-hui, ZHOU Yun-wei^*

College of Landscape Architecture, Northeast Forestry University, Harbin 150000, China

Received:2017-07-24 Revised:2017-10-18 Online:2018-02-20 Published:2018-02-20

摘要/Abstract

摘要： 从细叶百合cDNA文库中克隆得到ArfGAP基因,该基因ORF序列编码640个氨基酸,具有一个典型的C₄型ArfGAP保守结构域。蛋白相对分子量约为69.43 kDa,理论等电点为8.95,为亲水性蛋白,没有跨膜结构。氨基酸序列与油棕的GTPase同源性达到59%,与芭蕉、海枣、石刁柏、凤梨的同源性都在50%以上,命名为LpAGD14。qRT-PCR分析LpAGD14表达量呈不断升高的趋势,其中休眠完全解除时基因表达量显著高于其他时期。亚细胞定位显示该基因定位在细胞膜上。超微结构观察发现随低温贮藏时间的延长高尔基体囊泡层数逐渐增多。这些结果表明LpAGD14可能通过影响囊泡运输参与细叶百合鳞茎休眠解除进程。该研究为深入了解该基因功能奠定了基础。

Abstract: A gene encoding an ArfGAP protein was cloned from the cDNA library of Lilium pumilum. Its ORF sequence encoded a 640-amino acid polypeptide with a typical C4 ArfGAP conserved domain. The relative molecular mass of the putative ArfGAP protein was 69.43 kDa, the theoretical isoelectric point was 8.95, and it was predicted to be a hydrophilic protein with no transmembrane structure. At the amino acid sequence level, the L. pumilum ArfGAP showed 59% homology to ArfGAP-domain proteins (AGDs) in Elaeis guineensis, and more than 50% homology to AGDs in Musa acuminata subsp., Phoenix dactylifera, and Ananas comosus. Therefore, the gene was named LpAGD14. Transcript analysis by qRT-PCR showed that the transcript levels of LpAGD14 increased to reach the highest value after dormancy release. Subcellular localization assays showed that LpAGD14 localized to the cell membrane. Ultrastructural observations showed that the layers of the Golgi body increased with prolonged cold storage. These results indicated that LpAGD14 is involved in the dormancy release of L. pumilum and plays a role in vesicle transport. This study lays the foundation for further research on the function and regulation of this gene and its product.

汪王, 田忠平, 苏小霞, 杨柳慧, 周蕴薇. 细叶百合LpAGD14基因的克隆与表达分析[J]. 草业学报, 2018, 27(2): 117-123.

WANG Wang, TIAN Zhong-ping, SU Xiao-xia, YANG Liu-hui, ZHOU Yun-wei. Cloning and expression analysis of LpAGD14 from Lilium pumilum[J]. Acta Prataculturae Sinica, 2018, 27(2): 117-123.

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细叶百合LpAGD14基因的克隆与表达分析

Cloning and expression analysis of LpAGD14 from Lilium pumilum

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