金鱼草XTH家族基因鉴定及抗核盘菌和雄蕊瓣化相关基因筛选

doi:10.11983/CBB23107

植物学报 ›› 2024, Vol. 59 ›› Issue (2): 188-203.DOI: 10.11983/CBB23107

金鱼草XTH家族基因鉴定及抗核盘菌和雄蕊瓣化相关基因筛选

赵晗茜, 宋佳怡, 杨洁, 赵永晶, 夏文念, 顾伟卓, 汪仲毅, 杨楠, 胡慧贞^*()

西南林业大学园林园艺学院, 云南省功能性花卉资源及产业化技术工程中心, 昆明 650224

收稿日期:2023-08-07 接受日期:2024-01-30 出版日期:2024-03-01 发布日期:2024-01-30
通讯作者: * 胡慧贞, 云南省高层次人才引进计划青年人才, 国家林业和草原局荷花及水生植物产业研发创新联盟秘书长。2017年于华中农业大学获农学博士学位, 长期致力于荷花、金鱼草和油菜等植物切花和株型形成机理及细胞壁抗性研究。发表高质量SCI论文近20篇, 累计影响因子超过100; 主持国家级和省部级项目6项。E-mail: Jenny_8729@163.com
基金资助:
国家自然科学基金地区科学基金(32360070);国家自然科学基金青年基金(31901571);西南林业大学科研启动项目(01102-112109);云南省农业基础联合面上项目(202301BD070001-072)

Identification of XTH Family Genes in Antirrhinum majus and Screening of Genes Involoved in Sclerotinia sclerotiorum Resistance and Stamen Petalization

Hanqian Zhao, Jiayi Song, Jie Yang, Yongjing Zhao, Wennian Xia, Weizhuo Gu, Zhongyi Wang, Nan Yang, Huizhen Hu^*()

Yunnan Province Engineering Research Center for Functional Flower Resources and Industrialization, College of Landscape Architecture and Horticulture Sciences, Southwest Forestry University, Kunming 650224, China

Received:2023-08-07 Accepted:2024-01-30 Online:2024-03-01 Published:2024-01-30
Contact: * E-mail: Jenny_8729@163.com

摘要/Abstract

摘要： 木葡聚糖内转糖苷酶/水解酶(XTH)属于糖苷水解酶16家族(GH16), 是一类介导木葡聚糖(XyG)-纤维素骨架构建和重组的酶。为探明XTH家族基因在金鱼草(Antirrhinum majus)中的潜在生物学功能, 通过生物信息学分析, 结合转录组测序(RNA-seq)和实时荧光定量聚合酶链式反应(qRT-PCR)探究了XTH家族基因分别在金鱼草瓣化和非瓣化雄蕊以及抗感核盘菌材料中的表达水平。结果表明, 鉴定出的33个AmXTH蛋白主要保守基序为ExDxE, 分为3个亚组。AmXTH基因启动子的顺式作用元件多为生长发育、抗病及抗逆类。经RNA-seq和qRT-PCR验证, 最终挖掘出4个正向介导抗病的关键候选基因(AmXTH3、14、18和33), 1个负向介导抗病的关键候选基因(AmXTH23), 12个正向介导雄蕊瓣化的关键候选基因(AmXTH1、7、9、11、21、22、23、24、26、28、29和33)以及2个负向介导雄蕊瓣化的关键候选基因(AmXTH15和31); 其中AmXTH23 和AmXTH33可能同时在金鱼草抗核盘菌和雄蕊瓣化中发挥作用。该研究初步挖掘出参与金鱼草抗核盘菌及雄蕊瓣化的AmXTH候选基因, 为进一步揭示其生物学功能奠定了基础。

关键词: 金鱼草, 木葡聚糖内转糖苷酶/水解酶家族, 核盘菌抗性, 雄蕊瓣化, 基因差异表达

Abstract: Xyloglucan endotransglucosylase/hydrolases (XTH) belongs to glycoside hydrolase, family 16 (GH16) and it is a class of enzymes that mediate the construction and recombination of xyglucan-cellulose skeleton. To explore the potential biological functions of XTH family genes in Antirrhinum majus. In this paper, bioinformatics analysis, RNA-seq analysis and qRT-PCR were used to investigate the expression levels of the family genes in the petalized and non-petalized stamens and disease-resistant materials. The results showed that the main conserved motif from 33 identified AmXTH proteins was ExDxE, which could be divided into 3 subgroups. Most of the cis-acting elements of AmXTH promoter are growth and development, disease resistance and stress resistance. RNA-seq and qRT-PCR revalidation finally unearthed four positive candidate genes (AmXTH3, 14, 18, and 33) and one candidate genes (AmXTH23) for nagatively mediated Sclerotinia sclerotiorum resistance. There were 12 positive AmXTH candidate genes (AmXTH1, 7, 9, 11, 21, 22, 23, 24, 26, 28, 29 and 33) and 2 negative AmXTH candidate genes (AmXTH15 and 31). Among them, AmXTH23 and 33 may play a role in both of the A. majus resistant to S. sclerotiorum and stamen petalization. In this study, the candidate AmXTH genes involved in S. sclerotiorum resistance and stamen petalization of A. majus were preliminarily excavated, which laid a foundation for further revealing the gene function.

Key words: Antirrhinum majus, xyloglucan endotransglucosylase/hydrolases family, Sclerotinia sclerotiorum resistance, stamen petalization, differential gene expression

赵晗茜, 宋佳怡, 杨洁, 赵永晶, 夏文念, 顾伟卓, 汪仲毅, 杨楠, 胡慧贞. 金鱼草XTH家族基因鉴定及抗核盘菌和雄蕊瓣化相关基因筛选. 植物学报, 2024, 59(2): 188-203.

Hanqian Zhao, Jiayi Song, Jie Yang, Yongjing Zhao, Wennian Xia, Weizhuo Gu, Zhongyi Wang, Nan Yang, Huizhen Hu. Identification of XTH Family Genes in Antirrhinum majus and Screening of Genes Involoved in Sclerotinia sclerotiorum Resistance and Stamen Petalization. Chinese Bulletin of Botany, 2024, 59(2): 188-203.

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https://www.chinbullbotany.com/CN/Y2024/V59/I2/188

图/表 10

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Gene name	Forward primer (5′-3′)	Reverse primer (5′-3′)
AmUBI	ATCCACCCTTCACCTTGTG	TTGTCAATGGTATCCGAGC
AmXTH1	AAGGACAGAGGCAATGGGTG	AACCTGAGAACTCGGCAGTG
AmXTH2	TATGGAACGGAACGGGTTGG	TGCCAATTGTTTGGTGCGTT
AmXTH3	CGCCAAGATCTTCCGAGGAG	CAGCGACGAGTTTGAGTTGC
AmXTH4	AAAAGACCAAGCAATGGGCG	CTGCATCGTCGCATGTTGTC
AmXTH5	TATGGTTCGATCCCACTGCG	CGAACGGTGCCTTTTTCCAG
AmXTH6	GTGTTTGTGAACGGGCAAGG	TGCCCAACCATCTCCATTCC
AmXTH7	TGGTTTGATCCAACGGCTGA	TCCGTTGCCCAACTACTTCC
AmXTH8	TGCAGCTCTCAATCTGCCAA	GTAAAACGCGACGACGATGC
AmXTH9	GGCCGAATGCTTTCGCTATC	AGTTCCGGCTGAGTTTCCTG
AmXTH10	TCGCATTCCCGACTAACCAG	TACACGAATTCGAGAGCCCG
AmXTH11	ACGAAAGCTCCGTTTGTTGC	TCTCCCATTGGCGTGAAGTC
AmXTH12	GGCCAAATGCTGTCGCTATC	CAGTTCCGGCTGAGTTTCCT
AmXTH13	TACATCGGGCTCTGGTTTCG	ATCGTGTGCGTCTCCTTGAG
AmXTH14	CCCAAAAGCCAGCCAATGAG	GGCTTCGCAAAAGTGCAAGT
AmXTH15	CCACCGAGGAATCACTCCAG	TCTAGATTACGTCGTGCCGC
AmXTH16	TGCCGATGATTGGGCTACAC	TCAGGGGAAAGTTCACAGGC
AmXTH17	CAGGCTCCCTTTACCGCTTA	CGGGGGTGAGATCCCAATAC
AmXTH18	AGAACCACGACGAACTGGAC	AGGATGCTGTAACGATGCGA
AmXTH19	ACCTTCCCCTGGCTACTACC	TTCCTGAGGTTCTGTCGAGC
AmXTH20	TTCCTTGGCAATGTACCGGC	GCTGATCTGCAAGCTCATGG
AmXTH21	CCTCTGGAATCCTCAACGCA	CACTAGTCCGCCTCTTGTCG
AmXTH22	CGCTGAGTCAGTAGGTGTGG	TTTTCACTAGCCCGCCTCTC
AmXTH23	AAGTTCTGCGACACTCAGGG	TAGTGTAGCTGCTGCGAACC
AmXTH24	GTGGTTCGACCCGTCTAAGG	CTCCCCATTGCATCAACCCT
AmXTH25	GATGGATCAGAGTGGGCGAC	CCCACTGCAACCCAGTAGTC
AmXTH26	TGGGGCTATTTGAGCACTCG	GCCGAATGTGATTGTCAGCC
AmXTH27	TGGAACCCAATGGCTTGGAC	GGGTTGAGTTGCAAACTGGG
AmXTH28	CGACCCCTCCGAAGATTTCC	GCTATCCGCGTTCCAAATGC
AmXTH29	CGATTCTCACATTCGGCAGC	AGCTCGTCGCGTACTGTATC
AmXTH30	GAGCGGTTGTTCCGCATTTG	GCGTGAGAGCATGGTTCCTT
AmXTH31	TTTCGACATCACGTGGGGAG	TCCCAGGGACAAGCTTCAAC
AmXTH32	TCTACCGAGACAAATGGCCG	CAAGCGCATCTGGAAACGAG
AmXTH33	ATGAAGATATTTATTGTGCA	AGATACCAATGGGAATCTTC

Gene name	Forward primer (5′-3′)	Reverse primer (5′-3′)
AmUBI	ATCCACCCTTCACCTTGTG	TTGTCAATGGTATCCGAGC
AmXTH1	AAGGACAGAGGCAATGGGTG	AACCTGAGAACTCGGCAGTG
AmXTH2	TATGGAACGGAACGGGTTGG	TGCCAATTGTTTGGTGCGTT
AmXTH3	CGCCAAGATCTTCCGAGGAG	CAGCGACGAGTTTGAGTTGC
AmXTH4	AAAAGACCAAGCAATGGGCG	CTGCATCGTCGCATGTTGTC
AmXTH5	TATGGTTCGATCCCACTGCG	CGAACGGTGCCTTTTTCCAG
AmXTH6	GTGTTTGTGAACGGGCAAGG	TGCCCAACCATCTCCATTCC
AmXTH7	TGGTTTGATCCAACGGCTGA	TCCGTTGCCCAACTACTTCC
AmXTH8	TGCAGCTCTCAATCTGCCAA	GTAAAACGCGACGACGATGC
AmXTH9	GGCCGAATGCTTTCGCTATC	AGTTCCGGCTGAGTTTCCTG
AmXTH10	TCGCATTCCCGACTAACCAG	TACACGAATTCGAGAGCCCG
AmXTH11	ACGAAAGCTCCGTTTGTTGC	TCTCCCATTGGCGTGAAGTC
AmXTH12	GGCCAAATGCTGTCGCTATC	CAGTTCCGGCTGAGTTTCCT
AmXTH13	TACATCGGGCTCTGGTTTCG	ATCGTGTGCGTCTCCTTGAG
AmXTH14	CCCAAAAGCCAGCCAATGAG	GGCTTCGCAAAAGTGCAAGT
AmXTH15	CCACCGAGGAATCACTCCAG	TCTAGATTACGTCGTGCCGC
AmXTH16	TGCCGATGATTGGGCTACAC	TCAGGGGAAAGTTCACAGGC
AmXTH17	CAGGCTCCCTTTACCGCTTA	CGGGGGTGAGATCCCAATAC
AmXTH18	AGAACCACGACGAACTGGAC	AGGATGCTGTAACGATGCGA
AmXTH19	ACCTTCCCCTGGCTACTACC	TTCCTGAGGTTCTGTCGAGC
AmXTH20	TTCCTTGGCAATGTACCGGC	GCTGATCTGCAAGCTCATGG
AmXTH21	CCTCTGGAATCCTCAACGCA	CACTAGTCCGCCTCTTGTCG
AmXTH22	CGCTGAGTCAGTAGGTGTGG	TTTTCACTAGCCCGCCTCTC
AmXTH23	AAGTTCTGCGACACTCAGGG	TAGTGTAGCTGCTGCGAACC
AmXTH24	GTGGTTCGACCCGTCTAAGG	CTCCCCATTGCATCAACCCT
AmXTH25	GATGGATCAGAGTGGGCGAC	CCCACTGCAACCCAGTAGTC
AmXTH26	TGGGGCTATTTGAGCACTCG	GCCGAATGTGATTGTCAGCC
AmXTH27	TGGAACCCAATGGCTTGGAC	GGGTTGAGTTGCAAACTGGG
AmXTH28	CGACCCCTCCGAAGATTTCC	GCTATCCGCGTTCCAAATGC
AmXTH29	CGATTCTCACATTCGGCAGC	AGCTCGTCGCGTACTGTATC
AmXTH30	GAGCGGTTGTTCCGCATTTG	GCGTGAGAGCATGGTTCCTT
AmXTH31	TTTCGACATCACGTGGGGAG	TCCCAGGGACAAGCTTCAAC
AmXTH32	TCTACCGAGACAAATGGCCG	CAAGCGCATCTGGAAACGAG
AmXTH33	ATGAAGATATTTATTGTGCA	AGATACCAATGGGAATCTTC

Gene name	Gene ID	aa	Mw (Da)	PI	Instability index	Aliphatic index	GRAVY	TMHs	SP	Subcellular localization
AmXTH1	Am01g06240.T01	332	38077.94	6.37	43.80	71.30	-0.386	1	0	Cell wall
AmXTH2	Am01g11700.T01	260	29353.74	4.69	35.11	69.35	-0.338	0	1	Cell wall
AmXTH3	Am01g14580.T01	285	32755.11	9.27	38.06	72.53	-0.351	1	1	Cell wall, cytoplasmic
AmXTH4	Am01g53840.T01	290	33175.18	5.71	37.36	64.83	-0.420	0	1	Cell wall
AmXTH5	Am02g07210.T01	299	34771.82	5.28	34.83	58.70	-0.548	1	1	Cell wall
AmXTH6	Am02g15330.T01	221	25980.24	8.95	43.60	61.76	-0.759	0	0	Cell wall
AmXTH7	Am02g15340.T01	292	33675.55	6.13	24.95	64.38	-0.576	0	1	Cell wall
AmXTH8	Am02g39070.T01	340	39403.82	8.40	34.62	74.56	-0.463	1	1	Cell wall
AmXTH9	Am03g04070.T01	287	32230.44	8.56	34.96	74.46	-0.207	1	1	Cell wall
AmXTH10	Am03g04080.T01	287	32205.26	9.17	33.76	68.71	-0.290	0	1	Cell wall
AmXTH11	Am03g04460.T01	285	32026.84	8.21	31.88	64.04	-0.437	0	1	Cell wall
AmXTH12	Am03g04470.T01	278	31310.26	8.31	36.83	71.98	-0.216	0	1	Cell wall
AmXTH13	Am03g04480.T01	284	32223.01	4.82	36.01	69.75	-0.270	0	1	Cell wall
AmXTH14	Am03g08020.T01	288	32721.81	6.52	27.02	65.31	-0.416	0	1	Cell wall, cytoplasmic
AmXTH15	Am03g08030.T01	320	36873.73	8.63	30.08	70.41	-0.503	1	0	Cell wall
AmXTH16	Am04g01960.T01	303	35029.58	6.18	32.05	63.99	-0.456	1	1	Cell wall
AmXTH17	Am04g29790.T01	276	32119.17	5.00	47.65	66.81	-0.341	0	1	Cell wall
AmXTH18	Am05g24170.T01	350	40742.43	9.15	47.11	73.17	-0.491	0	0	Cell wall
AmXTH19	Am05g28140.T01	293	32825.66	7.08	40.82	65.60	-0.503	0	1	Cell wall
AmXTH20	Am05g34130.T01	294	34194.77	8.56	37.68	72.96	-0.397	1	1	Cell wall
AmXTH21	Am06g03100.T02	293	32816.60	5.12	40.48	65.97	-0.393	0	0	Cell wall, cytoplasmic
AmXTH22	Am06g03110.T02	294	32795.53	5.12	36.12	67.41	-0.404	1	1	Cell wall, cytoplasmic
AmXTH23	Am06g09100.T01	296	34193.50	8.22	48.24	68.21	-0.451	0	1	Cell wall, cytoplasmic
AmXTH24	Am06g11340.T01	304	35193.12	5.93	38.12	55.43	-0.663	0	1	Cell wall
AmXTH25	Am06g16890.T01	315	35592.11	7.14	51.07	73.59	-0.295	0	1	Cell wall
AmXTH26	Am06g38850.T01	288	32908.11	5.84	43.84	71.46	-0.362	1	1	Cell wall
AmXTH27	Am07g22440.T01	282	32198.49	8.70	39.91	74.65	-0.420	0	1	Cell wall
AmXTH28	Am07g32570.T01	290	33298.41	8.13	54.79	62.90	-0.449	0	1	Cell wall
AmXTH29	Am08g01110.T01	285	32630.86	6.38	35.18	69.12	-0.371	0	1	Cell wall
AmXTH30	Am08g26720.T01	298	34067.63	9.48	56.25	66.48	-0.351	1	1	Cell wall
AmXTH31	Am08g29760.T01	280	31431.47	5.93	39.93	68.64	-0.286	1	1	Cell wall, cytoplasmic
AmXTH32	Am08g29770.T01	647	73526.80	6.98	38.61	88.42	-0.058	1	0	Cell wall, cytoplasmic
AmXTH33	Am08g29790.T02	271	30595.34	6.41	31.72	63.32	-0.379	0	1	Cell wall, cytoplasmic

Gene name	Gene ID	aa	Mw (Da)	PI	Instability index	Aliphatic index	GRAVY	TMHs	SP	Subcellular localization
AmXTH1	Am01g06240.T01	332	38077.94	6.37	43.80	71.30	-0.386	1	0	Cell wall
AmXTH2	Am01g11700.T01	260	29353.74	4.69	35.11	69.35	-0.338	0	1	Cell wall
AmXTH3	Am01g14580.T01	285	32755.11	9.27	38.06	72.53	-0.351	1	1	Cell wall, cytoplasmic
AmXTH4	Am01g53840.T01	290	33175.18	5.71	37.36	64.83	-0.420	0	1	Cell wall
AmXTH5	Am02g07210.T01	299	34771.82	5.28	34.83	58.70	-0.548	1	1	Cell wall
AmXTH6	Am02g15330.T01	221	25980.24	8.95	43.60	61.76	-0.759	0	0	Cell wall
AmXTH7	Am02g15340.T01	292	33675.55	6.13	24.95	64.38	-0.576	0	1	Cell wall
AmXTH8	Am02g39070.T01	340	39403.82	8.40	34.62	74.56	-0.463	1	1	Cell wall
AmXTH9	Am03g04070.T01	287	32230.44	8.56	34.96	74.46	-0.207	1	1	Cell wall
AmXTH10	Am03g04080.T01	287	32205.26	9.17	33.76	68.71	-0.290	0	1	Cell wall
AmXTH11	Am03g04460.T01	285	32026.84	8.21	31.88	64.04	-0.437	0	1	Cell wall
AmXTH12	Am03g04470.T01	278	31310.26	8.31	36.83	71.98	-0.216	0	1	Cell wall
AmXTH13	Am03g04480.T01	284	32223.01	4.82	36.01	69.75	-0.270	0	1	Cell wall
AmXTH14	Am03g08020.T01	288	32721.81	6.52	27.02	65.31	-0.416	0	1	Cell wall, cytoplasmic
AmXTH15	Am03g08030.T01	320	36873.73	8.63	30.08	70.41	-0.503	1	0	Cell wall
AmXTH16	Am04g01960.T01	303	35029.58	6.18	32.05	63.99	-0.456	1	1	Cell wall
AmXTH17	Am04g29790.T01	276	32119.17	5.00	47.65	66.81	-0.341	0	1	Cell wall
AmXTH18	Am05g24170.T01	350	40742.43	9.15	47.11	73.17	-0.491	0	0	Cell wall
AmXTH19	Am05g28140.T01	293	32825.66	7.08	40.82	65.60	-0.503	0	1	Cell wall
AmXTH20	Am05g34130.T01	294	34194.77	8.56	37.68	72.96	-0.397	1	1	Cell wall
AmXTH21	Am06g03100.T02	293	32816.60	5.12	40.48	65.97	-0.393	0	0	Cell wall, cytoplasmic
AmXTH22	Am06g03110.T02	294	32795.53	5.12	36.12	67.41	-0.404	1	1	Cell wall, cytoplasmic
AmXTH23	Am06g09100.T01	296	34193.50	8.22	48.24	68.21	-0.451	0	1	Cell wall, cytoplasmic
AmXTH24	Am06g11340.T01	304	35193.12	5.93	38.12	55.43	-0.663	0	1	Cell wall
AmXTH25	Am06g16890.T01	315	35592.11	7.14	51.07	73.59	-0.295	0	1	Cell wall
AmXTH26	Am06g38850.T01	288	32908.11	5.84	43.84	71.46	-0.362	1	1	Cell wall
AmXTH27	Am07g22440.T01	282	32198.49	8.70	39.91	74.65	-0.420	0	1	Cell wall
AmXTH28	Am07g32570.T01	290	33298.41	8.13	54.79	62.90	-0.449	0	1	Cell wall
AmXTH29	Am08g01110.T01	285	32630.86	6.38	35.18	69.12	-0.371	0	1	Cell wall
AmXTH30	Am08g26720.T01	298	34067.63	9.48	56.25	66.48	-0.351	1	1	Cell wall
AmXTH31	Am08g29760.T01	280	31431.47	5.93	39.93	68.64	-0.286	1	1	Cell wall, cytoplasmic
AmXTH32	Am08g29770.T01	647	73526.80	6.98	38.61	88.42	-0.058	1	0	Cell wall, cytoplasmic
AmXTH33	Am08g29790.T02	271	30595.34	6.41	31.72	63.32	-0.379	0	1	Cell wall, cytoplasmic

金鱼草XTH家族基因鉴定及抗核盘菌和雄蕊瓣化相关基因筛选

Identification of XTH Family Genes in Antirrhinum majus and Screening of Genes Involoved in Sclerotinia sclerotiorum Resistance and Stamen Petalization

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