柑橘Rboh家族鉴定及其对激素和柑橘溃疡病的响应

doi:10.3864/j.issn.0578-1752.2020.20.008

Abstract

Abstract:

【Objective】Rboh is the main producer of reactive oxygen species (ROS), which is the key molecule involved in plant disease resistance signaling. To clarify the relationships between CsRboh family and citrus bacterial canker (CBC), the Rboh family of Citrus sinensis was identified and the bioinformatics characteristics as well as the expression patterns induced by biotic stress-related plant hormones and Xanthomonas citri subsp. citri (Xcc) were analyzed. It is also aimed to lay the foundation for further research of citrus Rboh genes in molecular breeding.【Method】Firstly, the RedOxiBase database and the citrus (sweet orange) CAP database were used to identify the citrus Rboh family sequences from the whole genome of C. sinensis. Then, series of bioinformatics tools were used to perform analysis of physical and chemical properties, phylogenetic relationships, chromosomal localizations, gene structures, protein functional domains, the conserved motifs and promoter elements systematically. Finally, qRT-PCR was used to analyze the expression patterns of CsRbohs induced by biotic stress-related plant hormones (salicylic acid, SA; jasmonic acid, JA; abscisic acid, ABA) and Xcc.【Result】A total of 7 members of the Rboh family in C. sinensis genome were identified (CsRboh01-CsRboh07). These genes encoded proteins contain 784-946 amino acid (AA) residues with isoelectric points (PI) ranging from 8.67 to 9.40. CsRbohs mainly distribute on the cell membrane system and cytoplasmic. The CsRbohs family can be divided into I to IV sub-groups based on the phylogenetic tree. These 7 genes are unevenly distributed on 5 sweet orange chromosomes. Each of the CsRbohs contains 4 typical functional domains, namely the respiratory burst NADPH oxidase domain (NADPH_Ox), the ferric reductase like transmembrane component (Ferric_reduct), the FAD binding domain (FAD_binding) and the ferric reductase NAD binding domain (NAD_binding). The promoter regions of CsRboh family genes contain different hormone response elements with different numbers and types. qRT-PCR showed that the expression of CsRboh gene family members in susceptible variety Wanjincheng and resistant variety Calamondin had some differences induced by different hormones and Xcc. According to the findings from phylogeny, homologous analysis and cis-acting elements analysis, CsRboh02, CsRboh04 and CsRboh06 had different expression patterns and levels induced by Xcc in resistant variety Calamondin and susceptible variety Wanjincheng.【Conclusion】CsRboh02, CsRboh04 and CsRboh06 may be closely related to the resistance and sensibility of citrus varieties, and they are three potential candidate genes for molecular breeding of resistance to canker disease, and the CsRboh family genes are preliminarily determined to play a key role in the process of citrus canker response.

Key words: citrus bacterial canker (CBC), Rboh family, Xanthomonas citri subsp. citri (Xcc), salicylic acid (SA), jasmonic acid (JA)

QIN XiuJuan,QI JingJing,DOU WanFu,CHEN ShanChun,HE YongRui,LI Qiang. Identification of Rboh Family and the Response to Hormone and Citrus Bacterial Canker in Citrus[J].Scientia Agricultura Sinica, 2020, 53(20): 4189-4203.

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References 60

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基因Gene	上游引物Forward primer	下游引物Reverse primer
CsRboh01	GCTCTCGGCTTCAACTTCCT	TCAAAGCTCGAGCCGCTAAA
CsRboh02	ATGGAGCACCAGCTCAAGAC	AGTAGCCCCAATTCCAAGCC
CsRboh03	TTGGCCAGCAGATTGGAGAG	CTGCGAAACTTGCCTCAACC
CsRboh04	ACGCCCGTCTTCAGATTTTCT	GCCATAAGTGACTGACACGCT
CsRboh05	CCGGCGCAATCTGTTAAAGG	TGTAGGCTTCAGCTTCTGGC
CsRboh06	CGCATGAAAGGGACACCGAAT	CAAATGAGCCTTGTTCCCTTGTT
CsRboh07	AACCGGTGATCAAGTGGTCG	GCATGGCTACAAAATTTGAGGGAA
CsActin	CATCCCTCAGCACCTTCC	CCAACCTTAGCACTTCTCC

名称 Name	CAP号 CAP ID	大小 Size (aa)	分子量 Molecular weight (Da)	等电点 Isoelectric point (PI)	亲水性平均值 Grand average of hydropathicity	脂肪指数 Aliphatic index	不稳定系数 Instability index	亚细胞定位 Subcellular localization
CsRboh01	Cs5g02940.1	946	106370.8	9.40	-0.247	87.68	50.16	细胞外膜Outer membrane
CsRboh02	Cs8g12000.1	912	103139.3	9.05	-0.299	83.84	42.57	细胞质Cytoplasmic
CsRboh03	Cs3g14240.1	889	101017.9	9.10	-0.265	85.43	40.96	细胞外膜Outer membrane
CsRboh04	Cs4g06920.1	875	99488.8	9.31	-0.252	82.61	48.38	细胞内膜Inner membrane
CsRboh05	Cs7g19320.1	811	91648.2	8.67	-0.269	86.94	40.91	细胞质Cytoplasmic
CsRboh06	Cs8g17640.1	844	96907.0	8.99	-0.173	84.69	40.42	细胞内膜Inner membrane
CsRboh07	Cs5g11890.1	784	89837.0	9.09	-0.015	89.06	44.93	细胞内膜Inner membrane

名称 Name	α-螺旋 Alpha helix (%)	β-转角 Beta turn (%)	不规则卷曲 Random coil (%)	伸展链 Extended strand (%)
CsRboh01	44.61	4.76	36.79	13.85
CsRboh02	43.75	5.26	34.87	16.12
CsRboh03	44.43	5.29	35.32	14.96
CsRboh04	41.83	4.57	38.29	15.31
CsRboh05	42.17	4.19	37.36	16.28
CsRboh06	43.60	5.21	36.85	14.34
CsRboh07	43.24	5.48	36.48	14.80

基因 Gene	茉莉酸响应元件 CGTCA-motif	水杨酸响应元件 TCA-element	脱落酸响应元件 ABRE
Csrboh01	3	1	7
Csrboh02	1	1	1
Csrboh03	2	-	5
Csrboh04	3	-	5
Csrboh05	-	-	6
Csrboh06 Csrboh07	2 1	1 -	1 -

Identification of Rboh Family and the Response to Hormone and Citrus Bacterial Canker in Citrus

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