马铃薯钼辅因子硫化酶基因(<i>LOS5</i>)克隆及功能分析

doi:10.7668/hbnxb.20196209

摘要/Abstract

摘要：

为探究马铃薯钼辅因子硫化酶基因(LOS5)的结构和功能,以紫彩3号马铃薯为试验材料,采用PCR技术克隆获得马铃薯StLOS5基因,并对其进行生物信息学分析、亚细胞定位、酵母逆境表型观察以及干旱胁迫下表达模式分析。结果表明,马铃薯StLOS5基因CDS区全长2 460 bp,编码819个氨基酸,分子质量为91.50 ku,理论等电点为6.78,亲水系数为-0.268,属于亲水性蛋白。系统进化分析显示,马铃薯StLOS5蛋白与番茄LOS5同源性最高,可能具有相似的功能。在其启动子区域检测到多个与非生物胁迫相关的顺式作用元件。亚细胞定位显示,StLOS5主要定位于细胞核。酵母逆境表型观察试验结果显示,StLOS5具有提高渗透胁迫耐受性的能力。荧光定量PCR分析结果表明,StLOS5的表达具有组织特异性,且在叶片中含量较高,并在干旱胁迫下能够快速响应,8%聚乙二醇处理12 h后叶片表达量达到峰值。以上结果表明,StLOS5在马铃薯响应干旱胁迫过程中具有一定作用。

关键词: 马铃薯, StLOS5, 生物信息学分析, 干旱胁迫

Abstract:

To investigate the structure and function of the potato molybdenum cofactor sulfurase(LOS5)gene,StLOS5 was cloned from the potato cultivar Zicai No.3 using PCR,followed by bioinformatics analysis,subcellular localization assays,yeast stress-tolerance assays,and expression pattern analysis under drought stress.The results showed that the CDS region of the potato StLOS5 gene was 2 460 bp in length,encoding 819 amino acids,with a molecular weight of 91.50 ku,a theoretical isoelectric point of 6.78,and a GRAVY value of -0.268,indicating that the protein was hydrophilic.Phylogenetic analysis revealed that the StLOS5 protein had the highest homology with tomato LOS5,suggesting that the two proteins may possess similar functions.Multiple cis-acting elements related to abiotic stress were detected in the promoter region.Subcellular localization showed that StLOS5 was primarily localized in the nucleus.Yeast stress-tolerance assays indicated that StLOS5 enhances tolerance to osmotic stress.Quantitative Real-time PCR analysis demonstrated that StLOS5 exhibits tissue-specific expression,with higher transcript levels in leaves,and responds rapidly to drought stress,with its expression in leaves peaking 12 h after treatment with 8%PEG.These results indicated that StLOS5 plays a role in the potato response to drought stress.

Key words: Potato, StLOS5, Bioinformatics analysis, Drought stress

中图分类号:

王英琪, 李景伟, 于卓, 赵晋邦, 袁清华, 刘博, 姜超. 马铃薯钼辅因子硫化酶基因(LOS5)克隆及功能分析[J]. 华北农学报, 2026, 41(1): 56-63. doi: 10.7668/hbnxb.20196209.

WANG Yingqi, LI Jingwei, YU Zhuo, ZHAO Jinbang, YUAN Qinghua, LIU Bo, JIANG Chao. Cloning and Functional Analysis of the Molybdenum Cofactor Sulfurase Gene(LOS5)in Potato[J]. Acta Agriculturae Boreali-Sinica, 2026, 41(1): 56-63. doi: 10.7668/hbnxb.20196209.

http://www.hbnxb.net/CN/Y2026/V41/I1/56

图/表 8

图1 StLOS5基因PCR电泳 M.DNA Marker 5000;1.PCR产物。

Fig.1 PCR electrophoretogram of StLOS5 gene M.DNA Marker 5000;1.PCR product.

图2 StLOS5蛋白磷酸化位点

Fig.2 Phosphorylation sites of StLOS5 protein

图3 StLOS5蛋白跨膜结构与信号肽

Fig.3 Transmembrane and signal peptide of StLOS5 protein

图4 StLOS5及其他同源蛋白的系统发育树

Fig.4 Phylogenetic tree of StLOS5 and other homologous proteins

表1 StLOS5启动子元件分析

Tab.1 Analysis of StLOS5 promoter elements

名称 Name	序列 Sequence	数量 Number	功能 Function
AAGAA-MOTIF	AAGAA	2	生长发育相关元件
ABRE	AGGTG	1	脱落酸响应元件
AC-Ⅱ	ACCAACC	1	脱落酸响应元件
ARE	TGGTTT	3	厌氧诱导元件
BOX-4	ATTAAT	3	光响应元件
CTAG-MOTIF	GATC	1	光响应元件
ERE	ATTTCAAA	3	乙烯响应元件
G-BOX	CACGTG	1	光响应元件
GC-MOTIF	CCCCCG	1	缺氧诱导元件
GT1-MOTIF	GGTTAA	2	光响应元件
W BOX	TTGAC	1	病原/防御相关元件
WRE3	CCACCT	2	损伤诱导元件
WUN-MOTIF	AAATTTCCT	1	创伤反应元件
LTR	CCGAA	1	低温响应元件
MYB	CAACTG	4	MYB类转录因子结合位点(主要参与干旱、盐分、ABA等胁迫应答)
MYB-LIKE SEQUENCE	TAACCA	1	MYB 类转录因子结合位点(光/激素应答模块)
MYC	CACATG	1	MYC 类转录因子结合位点(参与 ABA 介导的胁迫响应)
STRE	AGGGG	2	非特异性逆境响应元件
TATA-BOX	TATA	57	转录起始点核心启动子元件
TCA	TCAGAAGAGG	1	水杨酸响应元件
I-BOX	GATAAGG	1	光响应元件

图5 StLOS5蛋白在本氏烟草叶片中的亚细胞定位

Fig.5 Subcellular localization of StLOS5 protein in Nicotiana benthamiana leaves

图6 StLOS5酵母逆境表型观察

Fig.6 Yeast stress phenotyping of StLOS5

图7 8%聚乙二醇模拟干旱胁迫下StLOS5 基因表达分析不同小写字母表示组间差异显著(P<0.05)。

Fig.7 Expression analysis of StLOS5 gene under 8% PEG simulated drought stress Different letters indicate significant differences among treatments(P<0.05).

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马铃薯钼辅因子硫化酶基因(LOS5)克隆及功能分析

Cloning and Functional Analysis of the Molybdenum Cofactor Sulfurase Gene(LOS5)in Potato

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马铃薯钼辅因子硫化酶基因(LOS5)克隆及功能分析

Cloning and Functional Analysis of the Molybdenum Cofactor Sulfurase Gene(LOS5)in Potato

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