野生蒜芥茄<i>SsRPM1</i>基因的生信分析、亚细胞定位及表达分析

doi:10.7668/hbnxb.20195481

摘要/Abstract

摘要：

为了探究NBS-LRR类基因RPM1在茄属植物抗黄萎病中的作用,以茄属中的野生茄-蒜芥茄为材料,在其转录测序的基础上,克隆RPM1基因同源序列,分析该序列编码蛋白的理化性质、分子结构,构建进化树,以及在烟草中进行亚细胞定位,同时检测蒜芥茄不同部位中RPM1基因的相对表达量,以及在接种大丽轮枝菌(黄萎病病原菌的一种)后4个时间点(0,24,48,72 h)的相对表达量。结果发现,在蒜芥茄中RPM1基因(命名为SsRPM1)序列全长2 772 bp,编码 924 个氨基酸。SsRPM1蛋白总分子质量为105.99 ku,是不存在跨膜结构的碱性亲水蛋白,该蛋白主要由α螺旋和无规卷曲构成,包括LRR、NBC和CC 3种结构域,欧白英RPM1蛋白与其亲缘性关系最近;在烟草中的亚细胞定位发现该蛋白位于细胞膜上;SsRPM1基因在蒜芥茄的不同器官(根、茎、叶)中均有表达,其中茎的相对表达量最高,其次是叶,最后是根。在接种大丽轮枝菌后,总体上看,对照组和接菌处理组的SsRPM1基因表达情况皆呈现先上升后下降的趋势,在24 h最高。与对照组相比,接菌处理组的基因相对表达量较低,暗示蒜芥茄SsRPM1是响应黄萎病胁迫的负调控基因。

关键词: 野生茄, RPM1基因, 黄萎病, 生物信息学, 亚细胞定位, 基因表达

Abstract:

In order to explore the role of NBS-LRR gene RPM1 in the resistance to Verticillium wilt in Solanum,it took the wild Solanum sisymbriifolium Lam.as material,and cloned the homologous sequence of RPM1 gene on the basis of its transcriptional sequencing.The physicochemical properties and molecular structure of the sequence encoded protein were analyzed,the evolutionary relationship tree was constructed,and subcellular localization was performed in tobacco.At the same time,the relative expression level of RPM1 gene in different parts of Solanum sisymbriifolium Lam.was detected,as well as the relative expression level at for time points(0,24,48 and 72 h)after inoculation with Verticillium dahliae(a pathogen of Verticillium wilt).The results showed that the total length of RPM1 gene(SsRPM1)was 2 772 bp,encoding 924 amino acids.SsRPM1 protein,with a total molecular weight of 105.99 ku,was an alkaline hydrophilic protein without transmembrane structure.SsRPM1 protein was mainly composed of α helix and random coil,including LRR,NBC and CC domains.Solanum dulcamara RPM1 protein had the closest relationship with it.Subcellular localization in tobacco found that the protein was located on the cell membrane.SsRPM1 gene was expressed in different organs(root,stem and leaf),among which the stem had the highest relative expression,followed by leaf and root.After inoculation with V.dahliae,in general,SsRPM1 gene expression in both control group and inoculation group showed a trend of first increasing and then decreasing.The relative expression level of SsRPM1 gene was the highest at 24 h after inoculation.Compared with the control group,the relative expression level of SsRPM1 gene in inoculation treatment was lower.It is suggested that SsRPM1 is a negative regulatory gene in response to Verticillium wilt stress.

Key words: Wild eggplant, RPM1 gene, Verticillium wilt, Bioinformatics, Subcellular localization, Gene expression

中图分类号:

胡雨莹, 孙茂, 汪骞, 黎志彬, 鲍锐, 桂敏, 钟秋月, 杜光辉, 吴丽艳. 野生蒜芥茄SsRPM1基因的生信分析、亚细胞定位及表达分析[J]. 华北农学报, 2025, 40(3): 27-33. doi: 10.7668/hbnxb.20195481.

HU Yuying, SUN Mao, WANG Qian, LI Zhibin, BAO Rui, GUI Min, ZHONG Qiuyue, DU Guanghui, WU Liyan. Bioinformatics,Subcellular Localization and Expression Analysis of SsRPM1 Gene in Wild Solanum sisymbriifolium Lam.[J]. Acta Agriculturae Boreali-Sinica, 2025, 40(3): 27-33. doi: 10.7668/hbnxb.20195481.

http://www.hbnxb.net/CN/Y2025/V40/I3/27

图/表 10

图1 PCR扩增产物的1%琼脂糖凝胶电泳 M.5 000 bp DNA Marker;1—2.PCR产物。

Fig.1 1% agarose gel electrophoresis of PCR amplication products M.5 000 bp DNA Marker;1—2.PCR products.

图2 SsRPM1蛋白亲水性预测

Fig.2 Prediction result of hydrophobicity of SsRPM1 protein

图3 SsRPM1蛋白的二级结构(A)和三级(B)结构预测

Fig.3 Prediction of secondary structure(A) and tertiary structure(B) of SsRPM1 protein

图4 SsRPM1蛋白磷酸化位点预测

Fig.4 Prediction result of phosphoric acid sites in SsRPM1 protein

图5 SsRPM1蛋白结构域分析

Fig.5 Domain analysis of SsRPM1 protein

图6 5个物种中RPM1蛋白的氨基酸序列比对

Fig.6 Amino acid sequences alignment of RPM1 proteins in 5 different species

图7 不同植物RPM1蛋白的系统进化树

Fig.7 Phylogenetic tree of RPM1 proteins of different plant species

图8 SsRPM1蛋白在烟草细胞中亚细胞定位

Fig.8 Subcellular localization analysis in tobacco cells of SsRPM1 protein

图9 SsRPM1基因在蒜芥茄各器官中的相对表达量不同字母表示不同器官之间差异显著(P<0.05)。

Fig.9 Relative expression level of SsRPM1 gene in different organs of S.sisymbriifolium Different letters indicate significant difference between different organs(P<0.05).

图10 SsRPM1基因在蒜芥茄接种大丽轮枝菌后的相对表达量不同字母表示所有处理和时间点的差异显著(P<0.05)。

Fig.10 Relative expression level of SsRPM1 gene after S.sisymbriifolium inoculation with V.dahliae Different letters indicate significant differences between all the treatments and times(P<0.05).

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野生蒜芥茄SsRPM1基因的生信分析、亚细胞定位及表达分析

Bioinformatics,Subcellular Localization and Expression Analysis of SsRPM1 Gene in Wild Solanum sisymbriifolium Lam.

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野生蒜芥茄SsRPM1基因的生信分析、亚细胞定位及表达分析

Bioinformatics,Subcellular Localization and Expression Analysis of SsRPM1 Gene in Wild Solanum sisymbriifolium Lam.

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