小豆Dirigent基因家族鉴定及锈菌侵染对不同成员表达的影响

doi:10.3724/SP.J.1006.2022.14189

作物学报 ›› 2022, Vol. 48 ›› Issue (11): 2774-2785.doi: 10.3724/SP.J.1006.2022.14189

• 作物遗传育种·种质资源·分子遗传学 • 上一篇下一篇

小豆Dirigent基因家族鉴定及锈菌侵染对不同成员表达的影响

柯希望(), 苑梦琦(), 徐晓丹, 殷丽华, 郭永霞, 左豫虎^*()

黑龙江八一农垦大学农学院 / 黑龙江省作物有害生物互作生物学及生态防控重点实验室 / 国家杂粮工程技术研究中心,黑龙江大庆 163319

收稿日期:2021-10-18 接受日期:2022-02-25 出版日期:2022-11-12 网络出版日期:2022-03-23
通讯作者: 左豫虎
作者简介:第一作者联系方式: 柯希望, E-mail: kexylh@163.com
苑梦琦: E-mail: 2896834372@qq.com第一联系人：** 同等贡献。
基金资助:
本研究由国家自然科学基金项目(32102173);黑龙江省自然科学基金项目(YQ2020C034);黑龙江八一农垦大学研究生创新项目(YJSCX2021-Y50)

Dirigent gene family identification and expression profiling of Vigna angularis responsive to Uromyces vignae infection

KE Xi-Wang(), YUAN Meng-Qi(), XU Xiao-Dan, YIN Li-Hua, GUO Yong-Xia, ZUO Yu-Hu^*()

College of Agriculture, Heilongjiang Bayi Agricultural University / Heilongjiang Provincial Key Laboratory of Crop-Pest Interaction Biology Ecological Control / National Coarse Cereals Engineering Research Center, Daqing 163319, Heilongjiang, China

Received:2021-10-18 Accepted:2022-02-25 Published:2022-11-12 Published online:2022-03-23
Contact: ZUO Yu-Hu
Supported by:
The National Natural Science Foundation of China(32102173);The Heilongjiang Provincial Natural Science Foundation of China(YQ2020C034);The Innovative Research Program for Graduates of Heilongjiang Bayi Agricultural University(YJSCX2021-Y50)

摘要/Abstract

摘要：

为明确Dirigent (DIR)基因在小豆抗锈病中的作用,采用生物信息学方法,对小豆DIR基因家族(VaDIRs)进行了全基因组鉴定,结果发现小豆中共有33个DIR基因,其中29个成员分别定位在8条染色体上,系统进化分析表明,VaDIRs包括DIR-a、DIR-b/d、DIR-e和DIR-f四个亚家族,对VaDIRs成员启动子区顺式作用元件的分析发现,VaDIRs启动子区均包含激素、病原体应答等元件。对小豆抗锈病品种接种后不同时间的转录组数据分析发现,差异表达基因中共有17个VaDIRs成员,其中6个于接种后24 h显著上调,2个于接种后24 h和48 h均高量表达。进一步应用qRT-PCR技术对上述8个基因在抗、感品种中应答锈菌侵染的表达分析表明,高抗品种中VaDIR14、VaDIR16和VaDIR33的表达量在锈菌侵染不同阶段均显著高于感病品种。上述结果表明,VaDIRs家族成员可作为正调节因子参与小豆对锈病的抗性。

关键词: 小豆(Vigna angularis), 小豆锈病, 豇豆单胞锈菌(Uromyces vignae), DIR基因, 基因家族

Abstract:

To clarify the role of Dirigent (DIR) gene family of adzuki bean (Vigna angularis) in response to Uromyces vignae infection, genome-wide identification of the DIR family genes of V. angularis (VaDIRs) was conducted. A total of 33 VaDIRs in the V. angularis genome were identified, among which 29 members were located on 8 chromosomes, respectively. Phylogenetic analysis showed that VaDIRs consisted of 4 subfamilies (DIR-a, DIR-b/d, DIR-e, and DIR-f). Cis-acting elements analysis of the promoter of VaDIRs indicated that all members of VaDIRs contained hormones and pathogen responsive elements. Transcriptomic data of the rust resistant cultivar at different times after inoculation revealed that 17 VaDIRs members were differentially expressed, among which 6 VaDIRs were significantly upregulated at 24 hours post inoculation (hpi), and 2 members were highly expressed both at 24 and 48 hpi. The relative expression of the genes mentioned above were further confirmed in different resistant cultivars responsive to U. vignae infection by qRT-PCR. Results demonstrated that VaDIR14, VaDIR16, and VaDIR33 in resistant cultivar were significantly induced than those in the susceptible cultivar during the fungal infection. In short, our results indicated that members of VaDIRs could be used as positive regulators in resistant cultivar to prevent U. vignae infection.

Key words: Vigna angularis, adzuki bean rust, Uromyces vignae, DIR gene, gene family

柯希望, 苑梦琦, 徐晓丹, 殷丽华, 郭永霞, 左豫虎. 小豆Dirigent基因家族鉴定及锈菌侵染对不同成员表达的影响[J]. 作物学报, 2022, 48(11): 2774-2785.

KE Xi-Wang, YUAN Meng-Qi, XU Xiao-Dan, YIN Li-Hua, GUO Yong-Xia, ZUO Yu-Hu. Dirigent gene family identification and expression profiling of Vigna angularis responsive to Uromyces vignae infection[J]. Acta Agronomica Sinica, 2022, 48(11): 2774-2785.

图/表 9

表1

小豆DIR基因家族成员信息"

基因名称 Gene name	蛋白序列号 Protein sequence	基因组注释 Definition	染色体 Chr.	分子量 Molecular weight (kD)	等电点 pI	氨基酸数量 Number of the amino acids (aa)
VaDIR1	XP_017433474.1	未知蛋白Uncharacterized protein	01	19.78	9.16	185
VaDIR2	XP_017433817.1	类抗病反应蛋白206 Disease resistance response protein 206-like	01	20.35	6.28	182
VaDIR3	XP_017434121.1	类引导蛋白11 Dirigent protein 11-like	01	19.98	6.05	180
VaDIR4	XP_017414154.1	类引导蛋白22 Dirigent protein 22-like	02	20.79	7.07	190
VaDIR5	XP_017414268.1	类引导蛋白16 Dirigent protein 16-like	02	25.10	4.87	245
VaDIR6	XP_017412750.1	类引导蛋白16 Dirigent protein 16-like	02	27.37	5.10	266
VaDIR7	XP_017417575.1	类引导蛋白24 Dirigent protein 24-like	03	45.28	4.74	431
VaDIR8	XP_017416488.1	类引导蛋白25 Dirigent protein 25-like	03	24.22	5.78	224
VaDIR9	XP_017417865.1	类引导蛋白22 Dirigent protein 22-like	03	21.15	9.49	194
VaDIR10	XP_017416739.1	类引导蛋白22 Dirigent protein 22-like	03	20.98	9.40		192
VaDIR11	XP_017417602.1	类引导蛋白22 Dirigent protein 22-like	03	20.47	9.01		188
VaDIR12	XP_017417783.1	类引导蛋白2 Dirigent protein 2-like	03	20.34	5.22		186
VaDIR13	XP_017417792.1	类引导蛋白2 Dirigent protein 2-like	03	20.34	5.22		186
VaDIR14	XP_017417756.1	类引导蛋白22 Dirigent protein 22-like	03	24.04	6.59		216
VaDIR15	XP_017417215.1	类引导蛋白22 Dirigent protein 22-like	03	24.02	6.37		216
VaDIR16	XP_017417998.1	类引导蛋白22 Dirigent protein 22-like	03	24.17	7.07		216
VaDIR17	XP_017424454.1	类引导蛋白21 Dirigent protein 21-like	05	20.53	5.67		186
VaDIR18	XP_017426991.1	类引导蛋白11 Dirigent protein 11-like	06	21.11	8.83		196
VaDIR19	XP_017432020.1	类引导蛋白19 Dirigent protein 19-like	08	14.76	6.72		134
VaDIR20	XP_017432805.1	类引导蛋白2 Dirigent protein 2-like	08	19.32	9.45		177
VaDIR21	XP_017432829.1	类引导蛋白2 Dirigent protein 2-like	08	18.78	8.09		175
VaDIR22	XP_017438130.1	类引导蛋白22 Dirigent protein 22-like	10	15.03	6.32		135
VaDIR23	XP_017441922.1	类抗病反应蛋白206 Disease resistance response protein 206-like	11	21.32	8.58		189
VaDIR24	XP_017441955.1	类抗病反应蛋白206 Disease resistance response protein 206-like	11	20.22	6.95		183
VaDIR25	XP_017441660.1	类引导蛋白22 Dirigent protein 22-like	11	21.49	9.32		192
VaDIR26	XP_017441835.1	类引导蛋白22 Dirigent protein 22-like	11	21.02	9.25		189
VaDIR27	XP_017442075.1	类引导蛋白22 Dirigent protein 22-like	11	21.17	9.30		192
VaDIR28	XP_017442076.1	类引导蛋白22 Dirigent protein 22-like	11	20.49	9.30		187
VaDIR29	XP_017441889.1	类引导蛋白22 Dirigent protein 22-like	11	20.43	6.05		188
VaDIR30	XP_017411398.1	类引导蛋白9 Dirigent protein 9-like	/	26.77	6.30		248
VaDIR31	XP_017411409.1	类引导蛋白10 Dirigent protein 10 like	/	31.22	4.88		296
VaDIR32	XP_017405347.1	类引导蛋白22 Dirigent protein 22-like	/	21.78	6.97		190
VaDIR33	XP_017411321.1	未知蛋白 Uncharacterized protein	/	18.71	6.28		173

表1

图1

图2

图3

图4

图5

图6

表2

图7

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基因 Gene	上游引物 Upstream primer (5°-3°)		下游引物 Downstream primers (5°-3°)		产物长度 Product length (bp)
VaDIR9	GGAAAGGCCCAAGGTGTTTA		GCGTGCTGCCATTGTATTTC		103
VaDIR12	CGTCACCGCCACTTACTATC		GTCGTGGAAGAAGAAGTGAAGA		91
VaDIR14	GCCACCTTCTACCAAACTATCT		GGAGGGTTTGTCGTTTGTTATG		105
VaDIR15	GACCCACTGACCATCGGACCTG		CTGCGAGCAAGAACAAAGCCAC		260
基因 Gene	上游引物 Upstream primer (5°-3°)		下游引物 Downstream primers (5°-3°)		产物长度 Product length (bp)
VaDIR16		TCTTGTTTCTGGCACCTTCTAC		GTCTGATCGTCAATGGGTTCTC		206
VaDIR18		CCCATCTCAGCCACAACAACCG		CACTTCCTTCCACGCCATTCAC		129
VaDIR28		AGGTGGTCGGAAGAATTGAG		GCTGCTGCCATTGTACTTTC		110
VaDIR33		ATGTTTCAGCAATCGGCGTTCA		CAAGACCACGAGCAAAGGCAAA		148
VaACT		CTAAGGCTAATCGTGAGAA		CGTAAATAGGAACCGTGT		165

小豆Dirigent基因家族鉴定及锈菌侵染对不同成员表达的影响

Dirigent gene family identification and expression profiling of Vigna angularis responsive to Uromyces vignae infection

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