酵母双杂交系统筛选马铃薯<em>StLURP1</em>基因的互作蛋白

doi:10.3969/j.issn.1674-7968.2019.06.003

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摘要寄生霜霉菌响应迟发上调基因1(late up-regulated in response to Hyaloperonospora parasitica 1, LURP1)在植物抵御致病菌寄生霜霉菌(Hyaloperonospora parasitica, Hp)的反应和抗逆中起到关键作用。为了阐明马铃薯(Solanum tuberosum)LURP1基因的生物学功能并明确其在蛋白互作网络中的作用,本研究利用同源重组的定向克隆方式构建了其诱饵载体pGBKT7-StLURP1,并通过酵母(Saccharomyces cerevisiae)双杂交系统筛选马铃薯cDNA文库。结果表明,双杂筛选的阳性率约为70%,最终获得88个阳性克隆,经测序鉴定出12种含完整开放阅读框的StLURP1互作蛋白,互作的真实性通过回转实验进行验证。对互作蛋白进行Blast比对和基因本体注释(Gene Ontology, GO)分析,结果显示,StLURP1与甘油磷酸二酯酶、富脯氨酸蛋白、多酚氧化酶、肌动蛋白解聚因子、重金属离子转运蛋白及DnaJ家族蛋白等多种蛋白相互作用,可能参与植物脂类代谢、生物与非生物胁迫响应、肌动蛋白解聚、重金属离子解毒及蛋白质折叠等生物过程。本研究为进一步研究StLURP1基因的生物学功能与作用提供了理论依据。

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	房宸曦
	车妍
	廖钰秋
	王一凡
	张宁
	司怀军

关键词 ：马铃薯, 寄生霜霉菌响应迟发上调基因1(LURP1), 酵母双杂交, 诱饵载体, 互作蛋白

Abstract：The late up-regulated in response to Hyaloperonospora parasitica gene 1(LURP1) plays a key role in the response against pathogenic oomycete H. parasitica and resistance to stress in plants. In order to elucidate the biological function of the potato (Solanum tuberosum) LURP1 gene and to clarify its position in the protein interaction network, the bait vector pGBKT7-StLURP1 was constructed by homologous recombination and used to screen the potato cDNA library via the yeast (Saccharomyces cerevisiae) two-hybrid (Y2H) system in this research. The result showed that the positive rate of Y2H library screening was about 70%, and finally 88 positive clones were yielded, which was corresponded to 12 StLURP1 interacting proteins with complete ORF. The authenticity of the interactions was verified by retest. Blast alignment and Gene ontology (GO) annotation analysis demonstrated that StLURP1 could interact with the proteins such as DnaJ family protein, oxidoreductase/transcriptional repressor, glycerophosphodiester phosphodiesterase (GPGD), polyphenoloxidase, proline-rich protein, heavy metal transport/detoxification domain-containing protein, actin depolymerizing factor (ADF), nonspecific serine/threonine or tyrosine protein kinase, et al. The result indicated that StLURP1 may be involved in multiple biological processes, including lipid metabolism, biotic and abiotic stress responses, actin depolymerizing, heavy metal ions transport, protein folding and other biological processes. The present study would provide a theoretical foundation for further studying of biological function and role of StLURP1 gene.

Key words： Potato Late up-regulated in response to Hyaloperonospora parasitica gene 1 (LURP1) Yeast two-hybrid Bait vector Interacting protein

收稿日期: 2019-02-08

ZTFLH:

S532

基金资助:国家自然科学基金(No. 31660416;No. 31860399)

通讯作者: hjsi@gsau.edu.cn

引用本文:

房宸曦, 车妍, 廖钰秋, 王一凡, 张宁, 司怀军. 酵母双杂交系统筛选马铃薯StLURP1基因的互作蛋白[J]. 农业生物技术学报, 2019, 27(6): 972-981.
FANG Chen-Xi, CHE Yan, LIAO Yu-Qiu, WANG Yi-Fan, ZHANG Ning, SI Huai-Jun. Screening of Potato (Solanum tuberosum) Proteins interacting with StLURP1 Gene by Yeast (Saccharomyces cerevisiae) Two-Hybrid System. 农业生物技术学报, 2019, 27(6): 972-981.

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http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2019.06.003 或 http://journal05.magtech.org.cn/Jwk_ny/CN/Y2019/V27/I6/972

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