84个苹果栽培品种对斑点落叶病的抗性评价和全基因组关联分析

doi:10.3864/j.issn.0578-1752.2022.18.011

摘要/Abstract

摘要：

【目的】斑点落叶病是中国苹果产区发生的主要病害之一,严重影响苹果的产量和品质。本研究旨在发掘具有高抗病性的苹果栽培品种和探寻调控斑点落叶病抗性的关键基因,为苹果品种改良提供科学依据。【方法】利用苹果斑点落叶病菌（Alternaria alternata f. sp. mali）对84份苹果栽培品种进行离体叶片接种鉴定,从病斑面积和病斑面积增长率两方面进行聚类分析,评价苹果栽培品种对斑点落叶病的抗性。用接种后叶片的病斑面积作为表型性状,以全基因组深度重测序获得的1 243 071个高质量SNP位点为遗传标记,采用EMMAX方法进行全基因组关联分析。【结果】84份苹果栽培品种接种后统计病斑面积发现,不同的苹果栽培品种在对斑点落叶病的抗病性方面表现出显著的多样性,其中感病和中抗的品种占绝大多数,而高抗和高感的品种占比较少;苹果斑点落叶病抗病性具有正态分布特征,呈现数量性状遗传特征。全基因组关联性状分析最终获得6个SNP位点呈现显著水平P≤0.0000001（-LgP≥7）,深入分析将其关联到7个关键候选基因,包括整合素连接蛋白激酶、FMN连锁氧化还原酶、B-box型锌指蛋白、GATA型转录因子等,并验证了整合素连接蛋白激酶在苹果抗病中的作用。【结论】经过两年数据的综合分析,最终从84个苹果栽培品种里,鉴定到稳定抗性品种7份,稳定易感品种2份。通过全基因组关联性状分析鉴定到与苹果斑点落叶病抗病性显著相关的6个SNP位点,关联到7个关键候选基因,并验证了其中一个基因的功能。

关键词: 苹果栽培品种, 苹果斑点落叶病, 抗病性评价, 全基因组关联性状分析

Abstract:

【Objective】 Alternaria alternata f. sp. mali is a main disease occurring in the apple orchards in China, which seriously threatens the yield and quality of apples. The aim of this research was to identify apple cultivars and genes responsible for high resistance to Alternaria alternata f. sp. mali, thus providing the basis for disease resistance breeding of apples.【Method】 In this study, the isolated Alternaria alternata f. sp. mali strain was used to inoculate leaves of 84 apple cultivars. To evaluate the resistance of apple cultivars to Alternaria alternata f. sp. mali, the cluster analysis was carried out from two aspects including lesion area and lesion area growth rate. The genome-wide association study was performed based on the lesion area. The disease spot area of the inoculated leaves was used as a phenotypic trait, and the 1243071 high-quality SNP loci obtained by whole genome deep resequencing were used as genetic markers. Genome wide association analysis was performed by EMMAX method.【Result】The statistical analysis of lesion area showed significant diversity of 84 apple cultivars in resistance to Alternaria alternata f. sp. mali. Most apple cultivars were susceptible or resistant to Alternaria alternata f. sp. mali, while only a few cultivars were highly resistant and highly susceptible. The disease resistance of Alternaria alternata f. sp. mali has the characteristics of normal distribution, showing the genetic characteristics of quantitative characters. The genome-wide association trait analysis finally obtained 6 SNP loci with a significant level of P≤0.0000001 (-LgP≥7), and in-depth analysis linked them to 7 key candidate genes, including Integrin-linked protein kinase, FMN-linked oxidoreductase superfamily protein, B-box zinc finger family protein, GATA type transcription factor, etc. The biological function of Integrin-linked protein kinase in disease resistance was further verified. 【Conclusion】After two years of comprehensive analysis of data, 7 stable resistant varieties and 2 stable susceptible varieties were identified from 84 apple cultivars. Through genome-wide association trait analysis, six SNP loci and seven genes significantly related to the disease resistance of apple leaf spot were identified, and function of one gene was verified.

Key words: apple cultivars, Alternaria alternata f.sp. mali, disease resistance, genome-wide association study

储宝华,曹富国,卞宁宁,钱谦,李中兴,李雪薇,刘泽远,马锋旺,管清美. 84个苹果栽培品种对斑点落叶病的抗性评价和全基因组关联分析[J]. 中国农业科学, 2022, 55(18): 3613-3628.

BaoHua CHU,FuGuo CAO,NingNing BIAN,Qian QIAN,ZhongXing LI,XueWei LI,ZeYuan LIU,FengWang MA,QingMei GUAN. Resistant Evaluation of 84 Apple Cultivars to Alternaria alternata f. sp. mali and Genome-Wide Association Analysis[J]. Scientia Agricultura Sinica, 2022, 55(18): 3613-3628.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2022.18.011

https://www.chinaagrisci.com/CN/Y2022/V55/I18/3613

图/表 10

表1

表2

84个苹果属栽培品种离体叶片接种病斑大小及病斑面积增长率（2020年）"

品种名称 Cultivar resource	病斑面积 Lesion area (cm²)	增长率 Growth rate (%)	品种名称 Cultivar resource	病斑面积 Lesion area (cm²)	增长率 Growth rate (%)
凯密欧 Cameo	0.76±0.14	15.00	红露 Hongro	2.2±0.27	45.00
Kiku	0.90±0.17	15.75	斯派克 Spike	2.25±0.24	29.25
JA	0.95±0.21	19.50	平成 Heisei	2.31±0.19	48.75
皇家嘎啦 Royal Gala	0.99±0.13	16.75	阿肯色黑 Arkansasblack	2.38±0.24	50.25
秋光 Qiu Guang	1.01±0.33	19.50	布瑞本 Braeburn	2.4±0.18	53.25
蜜脆 Honeycrisp	1.03±0.32	19.50	Ce2	2.46±0.49	50.25
蓝皮尔曼 Blue Pearman	1.05±0.20	19.00	凉香 Ryoka	2.47±0.31	53.50
福拉瑞娜 Florina	1.14±0.33	19.00	Chenago Strawberry	2.49±0.17	54.75
Roho3615	1.20±0.14	23.25	秦阳 Qin Yang	2.51±0.21	50.25
艾达红 Idared	1.29±0.16	24.25	阿莱特 Arlet	2.51±0.32	55.00
威廉姆斯女士 Lady Williams	1.29±0.19	27.00	富红早嘎 Fu Hong Zao Ga	2.56±0.26	52.00
弘前富士 Hirosakifuji	1.33±0.17	22.25	Redcord	2.58±0.35	53.50
Fujiko	1.35±0.16	18.75	红勋1号 Hong Xun No.1	2.6±0.32	59.75
红盖露 Gale Gala	1.38±0.16	25.50	Ruby Mac	2.6±0.33	53.50
Gloster69	1.40±0.29	28.50	陆奥 Mutsu	2.61±0.34	60.00
长富2号 NaganofujiNo.2	1.42±0.27	26.25	秦冠 Qin Guan	2.69±0.32	60.75
早红1号 Early red one	1.45±0.20	25.00	Maririi Red	2.74±0.2	57.75
Challenger	1.61±0.18	29.50	千秋 Senshu	2.88±0.66	55.75
王林 Orin	1.62±0.30	31.00	新乔纳金 New Jonagold	2.88±0.91	61.50
鲁宾斯 Rubens	1.64±0.25	34.50	丽嘎 Li Ga	2.92±0.25	62.75
皮诺娃 Pinova	1.65±0.31	34.25	信浓甜 Cinano Sweet	2.93±0.65	61.00
N2	1.67±0.21	34.00	魔笛 Modi	3±0.36	66.50
无锈金冠 Reinders	1.68±0.36	35.00	富士冠军 Fuji Champion	3.05±0.06	62.00
坎兹 Kanzi	1.73±0.21	35.25	晨雾 Morning Mist	3.13±0.4	67.75
早红霞 Zao Hong Xia	1.74±0.32	30.25	金红 Jin Hong	3.14±0.27	63.25
玫瑰光芒 Rosy Glow	1.76±0.33	37.25	Northfield Beauty	3.17±0.21	69.50
美味 Ambrosia	1.77±0.18	36.25	北斗 Hokudo	3.26±0.98	66.75
宝罗红 PaulaRed	1.78±0.19	36.25	Yellow Transparent	3.29±0.68	67.50
Halerstadter Jungfermapfe	1.81±0.55	32.50	瑞林 Jude-line	3.37±0.08	71.75
北海道9号 HokkaidoNo.9	1.82±0.38	35.25	粉红女士 Pink Lady	3.42±0.16	70.25
夕阳 Sundowner	1.85±0.31	39.50	Su	3.45±0.3	65.50
巴克艾 Buckeye	1.86±0.15	35.00	K296	3.47±0.27	58.75
寒富 Han Fu	1.88±0.55	37.00	署红 James Grieve	3.64±0.29	68.00
荷斯坦 Holstein	1.90±0.12	39.75	茜 Akane	3.75±0.53	74.00
Unknown	1.90±0.25	41.75	Pound Sweet	3.83±0.16	80.75
瑞星 Judestar	1.92±0.23	36.00	2001富士 Fuji2001	4.12±0.47	81.25
昂林 Korin	1.93±0.21	34.25	阿丽亚娜 Ariane	4.24±0.49	80.75
瑞莲娜 Juliana	2.00±0.32	43.25	赫拉森 Haralson	4.32±0.68	75.50
津轻 Tsugaru	2.01±0.30	43.75	信浓红 CinanoRed	4.38±0.22	88.75
甘红 Gan Hong	2.07±0.33	38.00	比蒂格海姆 Bietigheimer	5.31±0.25	114.00
NJ-90	2.17±0.26	45.00	埃德尔博斯多夫 Edelborsdorfer	5.49±0.67	128.25
Ce1	2.19±0.27	47.00	Chanterler	6.02±0.23	95.75

表2

图1

表3

84个苹果属栽培品种离体叶片接种病斑大小及病斑面积增长率（2021年）"

品种名称 Cultivar resource	病斑面积 Lesion areas (cm²)	增长率 Growth rate (%)	品种名称 Cultivar resource	病斑面积 Lesion area (cm²)	增长率 Growth rate (%)
蓝皮尔曼 Blue Pearman	0.30±0.11	4.75	寒富 Han Fu	1.12±0.23	24.75
艾达红 Idared	0.36±0.19	8.00	坎兹 Kanzi	1.13±0.33	22.00
平成 Heisei	0.40±0.09	7.25	粉红女士 Pink Lady	1.17±0.35	22.25
Yellow Transparent	0.49±0.24	8.50	Maririi Red	1.18±0.23	26.25
Ce1	0.50±0.07	9.75	威廉姆斯女士 Lady Williams	1.20±0.28	23.50
皮诺娃 Pinova	0.60±0.27	13.00	瑞莲娜 Juliana	1.21±0.24	22.50
秦冠 Qin Guan	0.61±0.11	12.25	北海道9号 HokkaidoNo.9	1.21±0.25	22.50
NJ-90	0.62±0.22	11.50	红盖露 Gale Gala	1.23±0.26	25.00
无锈金冠 Reinders	0.63±0.10	12.00	2001富士 Fuji2001	1.23±0.34	26.75
宝罗红 PaulaRed	0.65±0.22	14.75	瑞林 Jude-line	1.23±0.36	25.75
皇家嘎啦 Royal Gala	0.65±0.33	13.00	巴克艾 Buckeye	1.30±0.12	25.25
玫瑰光芒 Rosy Glow	0.68±0.10	13.50	津轻 Tsugaru	1.33±0.27	28.25
夕阳 Sundowner	0.70±0.32	14.00	弘前富士 Hirosakifuji	1.37±0.25	30.75
Kiku	0.74±0.17	15.25	福拉瑞娜 Florina	1.37±0.29	28.25
信浓甜 Cinano Sweet	0.77±0.20	14.75	Chenago Strawberry	1.37±0.30	28.75
蜜脆 Honeycrisp	0.78±0.11	16.00	鲁宾斯 Rubens	1.37±0.32	27.50
阿莱特 Arlet	0.78±0.14	16.00	赫拉森 Haralson	1.41±0.10	22.75
Northfield Beauty	0.78±0.20	14.50	金红 Jin Hong	1.48±0.31	30.25
美味 Ambrosia	0.79±0.19	17.50	Unknown	1.50±0.61	30.25
凯密欧 Cameo	0.80±0.11	16.75	红勋1号 Hong Xun No.1	1.52±0.14	32.00
Challenger	0.80±0.13	16.50	魔笛 Modi	1.54±0.33	31.50
秦阳 Qin Yang	0.80±0.23	17.25	Pound Sweet	1.55±0.35	33.00
Su	0.81±0.28	18.25	早红1号 Early red one	1.55±0.43	33.25
Gloster69	0.82±0.17	15.50	比蒂格海姆 Bietigheimer	1.57±0.07	30.75
Ce2	0.83±0.20	17.75	K296	1.66±0.26	35.75
署红 James Grieve	0.87±0.10	17.25	北斗 Hokudo	1.69±0.34	31.25
凉香 Ryoka	0.90±0.25	19.75	N2	1.71±0.47	34.00
Fujiko	0.91±0.17	18.00	早红霞 Zao Hong Xia	1.75±0.16	37.00
丽嘎 Li Ga	0.92±0.20	18.50	斯派克 Spike	1.75±0.21	35.25
红露 Hongro	0.92±0.37	19.50	王林 Orin	1.80±0.41	37.75
陆奥 Mutsu	0.94±0.17	18.50	Redcord	1.80±0.73	38.25
秋光 Qiu Guang	0.94±0.21	17.75	埃德尔博斯多夫 Edelborsdorfer	1.84±0.2	34.25
富士冠军 Fuji Champion	0.95±0.40	19.00	昂林 Korin	1.86±0.07	36.00
瑞星 Judestar	0.96±0.15	19.25	富红早嘎 Fu Hong Zao Ga	1.87±0.16	41.00
布瑞本 Braeburn	0.99±0.13	17.75	甘红 Gan Hong	1.90±0.46	37.25
信浓红 CinanoRed	1.00±0.16	21.75	Chanterler	1.98±0.33	42.00
Roho3615	1.00±0.19	20.00	长富2号 NaganofujiNo.2	2.00±0.30	41.75
JA	1.01±0.18	20.75	新乔纳金 New Jonagold	2.05±0.44	42.75
阿肯色黑 Arkansasblack	1.03±0.21	20.25	千秋 Senshu	2.13±0.46	35.00
Halerstadter Jungfermapfe	1.08±0.18	20.00	Ruby Mac	2.14±0.49	44.75
晨雾 Morning Mist	1.10±0.25	23.50	荷斯坦 Holstein	2.37±0.48	44.00
阿丽亚娜 Ariane	1.12±0.20	24.00	茜 Akane	2.83±0.47	56.25

表3

图2

图3

表4

表5

图4

图5

参考文献 50

[1]	寿园园, 李春敏, 赵永波, 陈东玫, 张新忠, 杨国慧. 苹果早期落叶病的发生·防治及相关研究进展. 安徽农业科学, 2009, 37(20): 9519-9521. doi: 10.13989/j.cnki.0517-6611.2009.20.104. doi: 10.13989/j.cnki.0517-6611.2009.20.104
	SHOU Y Y, LI C M, ZHAO Y B, CHEN D M, ZHANG X Z, YANG G H. Occurrence, control and research development of the apple early stage leaf-cast. Journal of Anhui Agricultural Sciences, 2009, 37(20): 9519-9521. doi: 10.13989/j.cnki.0517-6611.2009.20.104. (in Chinese) doi: 10.13989/j.cnki.0517-6611.2009.20.104
[2]	ZHANG C X, TIAN Y, CONG P H. Proteome analysis of pathogen-responsive proteins from apple leaves induced by the Alternaria blotch Alternaria alternata. PLoS ONE, 2015, 10(6): e0122233. doi: 10.1371/journal.pone.0122233. doi: 10.1371/journal.pone.0122233
[3]	郑科. 苹果斑点落叶病的防治方法. 果农之友, 2020(5): 28.
	ZHENG K. Prevention and control method of apple speckled deciduous disease. Fruit Growers’ Friend, 2020(5): 28. (in Chinese)
[4]	赵国康, 李焰, 张树武, 徐秉良, 刘佳. 5种植物源农药对苹果斑点落叶病的防效评价. 中国果树, 2020(5): 46-49. doi: 10.16626/j.cnki.issn1000-8047.2020.05.008. doi: 10.16626/j.cnki.issn1000-8047.2020.05.008
	ZHAO G K, LI Y, ZHANG S W, XU B L, LIU J. Evaluation the control effects of five botanical fungicides on apple Alternaria leaf spot (Alternaria mali). China Fruits, 2020(5): 46-49. doi: 10.16626/j.cnki.issn1000-8047.2020.05.008. (in Chinese) doi: 10.16626/j.cnki.issn1000-8047.2020.05.008
[5]	郝丽霞, 张黎辉, 李畅, 刘希玲, 马俊欢. 38%唑醚∙戊唑醇悬浮剂对苹果树斑点落叶病的田间药效试验. 农药科学与管理, 2020, 41(3): 48-51.
	HAO L X, ZHANG L H, LI C, LIU X L, MA J H. Study on the control effect of pyraclostrobin + tebuconazole 38% SC against Myzus persicae. Pesticide Science and Administration, 2020, 41(3): 48-51. (in Chinese)
[6]	侯珲, 张恒涛, 周增强, 王丽, 阎振立, 王生荣. 苹果种质资源枝干轮纹病抗性评价. 园艺学报, 2017, 44(8): 1559-1568. doi: 10.16420/j.issn.0513-353x.2017-0286. doi: 10.16420/j.issn.0513-353x.2017-0286
	HOU H, ZHANG H T, ZHOU Z Q, WANG L, YAN Z L, WANG S R. Evaluation of resistance to apple ring rot in Malus germplasms. Acta Horticulturae Sinica, 2017, 44(8): 1559-1568. doi: 10.16420/j.issn.0513-353x.2017-0286. (in Chinese) doi: 10.16420/j.issn.0513-353x.2017-0286
[7]	张楠, 冯浩, 宋琳琳, 伏波, 高小宁, 黄丽丽. 苹果种质资源对苹果树腐烂病的抗性评价. 中国果树, 2019(3): 74-76, 80. doi: 10.16626/j.cnki.issn1000-8047.2019.03.018. doi: 10.16626/j.cnki.issn1000-8047.2019.03.018
	ZHANG N, FENG H, SONG L L, FU B, GAO X N, HUANG L L. Evaluation of resistance of apple germplasm resources to apple canker (Valsa mali). China Fruits, 2019(3): 74-76, 80. doi: 10.16626/j.cnki.issn1000-8047.2019.03.018. (in Chinese) doi: 10.16626/j.cnki.issn1000-8047.2019.03.018
[8]	YIN L H, WANG P, LI M J, KE X W, LI C Y, LIANG D, WU S, MA X L, LI C, ZOU Y J, MA F W. Exogenous melatonin improves Malus resistance to Marssonina apple blotch. Journal of Pineal Research, 2013, 54(4): 426-434. doi: 10.1111/jpi.12038. doi: 10.1111/jpi.12038
[9]	张坤, 党志国, 赵磊, 赵政阳. 富士、秦冠苹果对早期落叶病抗性的遗传分析. 西北林学院学报, 2007, 22(4): 128-130, 138. doi: 10.3969/j.issn.1001-7461.2007.04.032. doi: 10.3969/j.issn.1001-7461.2007.04.032
	ZHANG K, DANG Z G, ZHAO L, ZHAO Z Y. Study on inheritance tendency of the resistance to apple early defoliation diseases using ‘Qinguan’ and ‘fuji’. Journal of Northwest Forestry University, 2007, 22(4): 128-130, 138. doi: 10.3969/j.issn.1001-7461.2007.04.032. (in Chinese) doi: 10.3969/j.issn.1001-7461.2007.04.032
[10]	何晓燕. 苹果斑点落叶病的综合防治技术. 果农之友, 2013(9): 18. doi: 10.3969/j.issn.1671-7759.2013.09.016. doi: 10.3969/j.issn.1671-7759.2013.09.016
	HE X Y. Integrated control technology of apple speckled deciduous disease. Fruit Growers' Friend, 2013(9): 18. doi: 10.3969/j.issn.1671-7759.2013.09.016. (in Chinese) doi: 10.3969/j.issn.1671-7759.2013.09.016
[11]	任生林, 吴才文, 经艳芬, 刘家勇. 全基因组关联分析在作物中的研究进展. 分子植物育种, 2021. http://kns.cnki.net/kcms/detail/ 46.1068.S.20210929.1449.008.html.
	REN S L, WU W C, JING Y F, LIU J Y. Research progress of genome-wide association analysis in crops. Molecular Plant Breeding, 2021. http://kns.cnki.net/kcms/detail/ 46.1068.S.20210929.1449.008.html. (in Chinese)
[12]	PAULINO J F C, ALMEIDA C P, BUENO C J, SONG Q, FRITSCHE- NETO R, CARBONELL S A M, CHIORATO A F, BENCHIMOL- REIS L L. Genome-wide association study reveals genomic regions associated with Fusarium wilt resistance in common bean. Genes, 2021, 12(5): 765. doi: 10.3390/genes12050765. doi: 10.3390/genes12050765
[13]	YANG Y, AMO A, WEI D, CHAI Y M, ZHENG J, QIAO P F, CUI C G, LU S, CHEN L, HU Y G. Large-scale integration of meta-QTL and genome-wide association study discovers the genomic regions and candidate genes for yield and yield-related traits in bread wheat. Theoretical and Applied Genetics, 2021, 134(9): 3083-3109. doi: 10.1007/s00122-021-03881-4. doi: 10.1007/s00122-021-03881-4
[14]	FARNETI B, DI GUARDO M, KHOMENKO I, CAPPELLIN L, BIASIOLI F, VELASCO R, COSTA F. Genome-wide association study unravels the genetic control of the apple volatilome and its interplay with fruit texture. Journal of Experimental Botany, 2017, 68(7): 1467-1478. doi: 10.1093/jxb/erx018. doi: 10.1093/jxb/erx018
[15]	LIAO L, ZHANG W H, ZHANG B, FANG T, WANG X F, CAI Y M, OGUTU C, GAO L, CHEN G, NIE X Q, XU J S, ZHANG Q Y, REN Y R, YU J Q, WANG C K, DENG C H, MA B, ZHENG B B, YOU C X, HU D G, ESPLEY R, WANG K L, YAO J L, ALLAN A C, KHAN A, KORBAN S S, FEI Z J, MING R, HAO Y J, LI L, HAN Y P. Unraveling a genetic roadmap for improved taste in the domesticated apple. Molecular Plant, 2021, 14(9): 1454-1471. doi: 10.1016/j.molp.2021.05.018. doi: 10.1016/j.molp.2021.05.018
[16]	杨锋, 刘晨, 姜丽娟, 管清美. 苹果属植物抗旱性评价. 西北农林科技大学学报(自然科学版). 2020, 48(8): 119-128. doi: 10.13207/j.cnki.jnwafu.2020.08.015. doi: 10.13207/j.cnki.jnwafu.2020.08.015
	YANG F, LIU C, JIANG L J, GUAN Q M. Comprehensive evaluation on drought tolerance of Malus. Journal of Northwest A&F University (National Science Edition), 2020, 48(8): 119-128. doi: 10.13207/j.cnki.jnwafu.2020.08.015. (in Chinese) doi: 10.13207/j.cnki.jnwafu.2020.08.015
[17]	ZHANG Q L, MA C, ZHANG Y, GU Z Y, LI W, DUAN X W, WANG S N, HAO L, WANG Y H, WANG S Y, LI T Z. A single-nucleotide polymorphism in the promoter of a hairpin RNA contributes to Alternaria alternata leaf spot resistance in apple (Malus×domestica). The Plant Cell, 2018, 30(8): 1924-1942. doi: 10.1105/tpc.18.00042. doi: 10.1105/tpc.18.00042
[18]	CHEN P X, LI Z X, ZHANG D H, SHEN W Y, XIE Y P, ZHANG J, JIANG L J, LI X W, SHEN X X, GENG D L, WANG L P, NIU C D, BAO C N, YAN M J, LI H Y, LI C Y, YAN Y, ZOU Y J, MICHELETTI D, KOOT E, MA F W, GUAN Q M. Insights into the effect of human civilization on Malus evolution and domestication. Plant Biotechnology Journal, 2021, 19(11): 2206-2220. doi: 10.1111/pbi.13648. doi: 10.1111/pbi.13648
[19]	LI X W, ZHOU S X, LIU Z Y, LU L Y, DANG H, LI H M, CHU B H, CHEN P X, MA Z Q, ZHAO S, LI Z X, NOCKER S V, MA F W, GUAN Q M. Fine-tuning of SUMOylation modulates drought tolerance of apple, Plant Biotechnology Journal, 2022, 20(5): 903-919. DOI: 10.1111/pbi.13772. doi: 10.1111/pbi.13772
[20]	NEMOTO K, SETO T, TAKAHASHI H, NOZAWA A, SEKI M, SHINOZAKI K, ENDO Y, SAWASAKI T. Autophosphorylation profiling of Arabidopsis protein kinases using the cell-free system. Phytochemistry, 2011, 72(10): 1136-1144. doi: 10.1016/j.phytochem.2011.02.029. doi: 10.1016/j.phytochem.2011.02.029
[21]	ASCENCIO-IBÁÑEZ J T, SOZZANI R, LEE T J, CHU T M, WOLFINGER R D, CELLA R, HANLEY-BOWDOIN L. Global analysis of Arabidopsis gene expression uncovers a complex array of changes impacting pathogen response and cell cycle during geminivirus infection. Plant Physiology, 2008, 148(1): 436-454. doi: 10.1104/pp.108.121038. doi: 10.1104/pp.108.121038
[22]	WANG D, GUO Y H, WU C G, YANG G D, LI Y Y, ZHENG C C. Genome-wide analysis of CCCH zinc finger family in Arabidopsis and rice. BMC Genomics, 2008, 9 (1): 44. doi: 10.1186/1471-2164-9-44. doi: 10.1186/1471-2164-9-44
[23]	DAI H Y, LI W R, HAN G F, YANG Y, MA Y, LI H, ZHANG Z H. Development of a seedling clone with high regeneration capacity and susceptibility to Agrobacterium in apple. Scientia Horticulturae, 2013, 164: 202-208. doi: 10.1016/j.scienta.2013.09.033. doi: 10.1016/j.scienta.2013.09.033
[24]	常亚洲, 张会龙. 苹果早期落叶病的发生与防治. 中国果菜, 2011, 31(9): 31. doi: 10.3969/j.issn.1008-1038.2011.09.023. doi: 10.3969/j.issn.1008-1038.2011.09.023
	CHANG Y Z, ZHANG H L. Occurrence and control of apple deciduous disease at early stage. China Fruit and Vegetable, 2011, 31(9): 31. doi: 10.3969/j.issn.1008-1038.2011.09.023. (in Chinese) doi: 10.3969/j.issn.1008-1038.2011.09.023
[25]	郭小侠, 陈川, 唐周怀, 石晓红, 石勇强. 苹果早期落叶病的发生规律及生物防治. 陕西农业科学, 2004, 50(1): 62-64.
	GUO X X, CHEN C, TANG Z H, SHI X H, SHI Y Q. Occurrence regularity and biological control of apple deciduous disease in early stage. Shaanxi Journal of Agricultural Sciences, 2004, 50(1): 62-64. (in Chinese)
[26]	李爽, 张军科, 党伟锋. 苹果遗传连锁图谱构建及抗早期落叶病的基因定位. 北方园艺, 2011(24): 145-149.
	LI S, ZHANG J K, DANG W F. Genetic mapping and localization of the early deciduous disease resistant gene in apple. Northern Horticulture, 2011(24): 145-149. (in Chinese)
[27]	GENG D L, SHEN X X, XIE Y P, YANG Y S, BIAN R L, GAO Y Q, LI P M, SUN L Y, FENG H, MA F W, GUAN Q M. Regulation of phenylpropanoid biosynthesis by MdMYB88 and MdMYB124 contributes to pathogen and drought resistance in apple. Horticulture Research, 2020, 7: 102. doi: 10.1038/s41438-020-0324-2. doi: 10.1038/s41438-020-0324-2
[28]	李鹏, 王益权, 焦彩强, 石宗琳, 梁化学. 陕西渭北地区苹果主栽品种的品质分析与评价. 西北农业学报, 2016, 25(9): 1358-1364. doi: 10.7606/j.issn.1004-1389.2016.09.012. doi: 10.7606/j.issn.1004-1389.2016.09.012
	LI P, WANG Y Q, JIAO C Q, SHI Z L, LIANG H X. Fruit quality evaluation of three apple cultivars in north region of Weihe River in Shaanxi Province. Acta Agriculturae Boreali-Occidentalis Sinica, 2016, 25(9): 1358-1364. doi: 10.7606/j.issn.1004-1389.2016.09.012. (in Chinese) doi: 10.7606/j.issn.1004-1389.2016.09.012
[29]	刘振西, 韩立新, 霍振芳, 郝贝贝, 武少杰. 苹果品种“秦阳”在三门峡的引种表现. 河南农业, 2019(1): 45. doi: 10.15904/j.cnki.hnny.2019.01.037. doi: 10.15904/j.cnki.hnny.2019.01.037
	LIU Z X, HAN L X, HUO Z F, HAO B B, WU S J. Introduction of apple variety ‘Qinyang’ in Sanmenxia. Agriculture of Henan, 2019(1): 45. doi: 10.15904/j.cnki.hnny.2019.01.037. (in Chinese) doi: 10.15904/j.cnki.hnny.2019.01.037
[30]	周建林, 刘国成, 秦嗣军. ‘寒富’苹果无袋栽培主要病虫害的防控. 北方果树, 2021(3): 33-35. doi: 10.16376/j.cnki.bfgs.2021.03.011. doi: 10.16376/j.cnki.bfgs.2021.03.011
	ZHOU J L, LIU G C, QIN S J. Prevention and control of main diseases and insect pests in bagless cultivation of ‘Hanfu’ apple. Northern Fruits, 2021(3): 33-35. doi: 10.16376/j.cnki.bfgs.2021.03.011. (in Chinese) doi: 10.16376/j.cnki.bfgs.2021.03.011
[31]	张秉宇, 刘志. 不同砧穗组合对“寒富”苹果果实品质的影响. 北方园艺, 2014(7): 30-32.
	ZHANG B Y, LIU Z. Effect of different scion-sotck combinations on the quality of ‘hanfu’ apple. Northern Horticulture, 2014(7): 30-32. (in Chinese)
[32]	路新创, 陈新宝, 赵上利, 赵海文. 苹果中熟品种蜜脆引种体会. 西北园艺(果树), 2018(1): 33-34.
	LU X C, CHEN X B, ZHAO S L, ZHAO H W. Introduction of apple medium ripe variety honey crisp. Northwest Horticulture, 2018(1): 33-34. (in Chinese)
[33]	CHEN Q M, DONG C H, SUN X H, ZHANG Y G, DAI H Y, BAI S H. Overexpression of an apple LysM-containing protein gene, MdCERK1-2, confers improved resistance to the pathogenic fungus, Alternaria alternata, in Nicotiana benthamiana. BMC Plant Biology, 2020, 20(1): 146. doi: 10.1186/s12870-020-02361-z. doi: 10.1186/s12870-020-02361-z
[34]	HOU Y J, YU X Y, CHEN W P, ZHUANG W B, WANG S H, SUN C, CAO L F, ZHOU T T, QU S C. MdWRKY75e enhances resistance to Alternaria alternata in Malus domestica. Horticulture Research, 2021, 8: 225. doi: 10.1038/s41438-021-00701-0. doi: 10.1038/s41438-021-00701-0
[35]	ZHANG Q L, XU C R, WEI H Y, FAN W Q, LI T Z. Two pathogenesis-related proteins interact with leucine-rich repeat proteins to promote Alternaria leaf spot resistance in apple. Horticulture Research, 2021, 8: 219. doi: 10.1038/s41438-021-00654-4. doi: 10.1038/s41438-021-00654-4
[36]	HUANG K, ZHONG Y, LI Y, ZHENG D, CHENG Z M. Genome-wide identification and expression analysis of the apple ASR gene family in response to Alternaria alternata f.sp. Mali Genome, 2016, 59(10): 866-878. doi: 10.1139/gen-2016-0043. doi: 10.1139/gen-2016-0043
[37]	JI S D, LIU Z H, WANG Y C. Trichoderma-induced ethylene responsive factor MsERF105 mediates defense responses in Malus sieversii. Frontiers in Plant Science, 2021, 12: 708010. doi: 10.3389/fpls.2021.708010. doi: 10.3389/fpls.2021.708010
[38]	ASCENCIO-IBÁÑEZ J T, SOZZANI R, LEE T J, CHU T M, WOLFINGER R D, CELLA R, HANLEY-BOWDOIN L. Global analysis of Arabidopsis gene expression uncovers a complex array of changes impacting pathogen response and cell cycle during geminivirus infection. Plant Physiology, 2008, 148(1): 436-454. doi: 10.1104/pp.108.121038. doi: 10.1104/pp.108.121038
[39]	严勇亮, 时晓磊, 张金波, 耿洪伟, 肖菁, 路子峰, 倪中福, 丛花. 春小麦籽粒主要品质性状的全基因组关联分析. 中国农业科学, 2021, 54(19): 4033-4047. doi: 10.3864/j.issn.0578-1752.2021.19.001. doi: 10.3864/j.issn.0578-1752.2021.19.001
	YAN Y L, SHI X L, ZHANG J B, GENG H W, XIAO J, LU Z F, NI Z F, CONG H. Genome-wide association study of grain quality related characteristics of spring wheat. Scientia Agricultura Sinica, 2021, 54(19): 4033-4047. doi: 10.3864/j.issn.0578-1752.2021.19.001. (in Chinese) doi: 10.3864/j.issn.0578-1752.2021.19.001
[40]	于海飞, 杜晓宇, 殷贵鸿, 邹少奎, 李楠, 张倩, 吕永军, 王丽娜, 王雅美, 韩玉林. 普通小麦抗倒伏相关性状的全基因组关联分析. 植物遗传资源学报, 2022, 23(1): 147-159. doi: 10.13430/j.cnki.pngr.20210731001. doi: 10.13430/j.cnki.pngr.20210731001
	YU H F, DU X Y, YIN G H, ZOU S K, LI N, ZHANG Q, LÜ Y J, WANG L N, WANG Y M, HAN Y L. Genome-wide association mapping for lodging-resistance related traits in common wheat (Triticum aestivum L.). Journal of Plant Genetic Resources, 2022, 23(1): 147-159. doi: 10.13430/j.cnki.pngr.20210731001. (in Chinese) doi: 10.13430/j.cnki.pngr.20210731001
[41]	TAHERI P, TARIGHI S. A survey on basal resistance and riboflavin- induced defense responses of sugar beet against Rhizoctonia solani. Journal of Plant Physiology, 2011, 168(10): 1114-1122. doi: 10.1016/j.jplph.2011.01.001. doi: 10.1016/j.jplph.2011.01.001
[42]	DENG B L, DENG S, SUN F, ZHANG S J, DONG H S. Down-regulation of free riboflavin content induces hydrogen peroxide and a pathogen defense in Arabidopsis. Plant Molecular Biology, 2011, 77(1/2): 185-201. doi: 10.1007/s11103-011-9802-0. doi: 10.1007/s11103-011-9802-0
[43]	ZHANG S, YANG X, SUN M W, SUN F, DENG S, DONG H S. Riboflavin-induced priming for pathogen defense in Arabidopsis thaliana. Journal of Integrative Plant Biology, 2009, 51(2): 167-174. doi: 10.1111/j.1744-7909.2008.00763.x. doi: 10.1111/j.1744-7909.2008.00763.x.
[44]	BRAUER E K, AHSAN N, DALE R, KATO N, COLUCCIO A E, PIÑEROS M A, KOCHIAN L V, THELEN J J, POPESCU S C. The raf-like kinase ILK1 and the high affinity k⁺ transporter HAK5 are required for innate immunity and abiotic stress response. Plant Physiology, 2016, 171(2): 1470-1484. doi: 10.1104/pp.16.00035. doi: 10.1104/pp.16.00035
[45]	AMORIM L L B, DA FONSECA DOS SANTOS R, NETO J P B, GUIDA-SANTOS M, CROVELLA S, BENKO-ISEPPON A M. Transcription factors involved in plant resistance to pathogens. Current Protein & Peptide Science, 2017, 18(4): 335-351. doi: 10.2174/1389203717666160619185308. doi: 10.2174/1389203717666160619185308
[46]	袁岐, 张春利, 赵婷婷, 许向阳. 植物中GATA转录因子的研究进展. 分子植物育种, 2017, 15(5): 1702-1707. doi: 10.13271/j.mpb.015.001702. doi: 10.13271/j.mpb.015.001702
	YUAN Q, ZHANG C L, ZHAO T T, XU X Y. Research advances of GATA transcription factor in plant. Molecular Plant Breeding, 2017, 15(5): 1702-1707. doi: 10.13271/j.mpb.015.001702. (in Chinese) doi: 10.13271/j.mpb.015.001702
[47]	MOON S J, PARK H J, KIM T H, KANG J W, LEE J Y, CHO J H, LEE J H, PARK D S, BYUN M O, KIM B G, SHIN D. OsTGA2 confers disease resistance to rice against leaf blight by regulating expression levels of disease related genes via interaction with NH₁. PLoS ONE, 2018, 13(11): e0206910. doi: 10.1371/journal.pone.0206910. doi: 10.1371/journal.pone.0206910
[48]	LIM C W, BAEK W, LIM S, HAN S W, LEE S C. Expression and functional roles of the pepper pathogen-induced bZIP transcription factor CabZIP₂ in enhanced disease resistance to bacterial pathogen infection. Molecular Plant-Microbe Interactions, 2015, 28(7): 825-833. doi: 10.1094/mpmi-10-14-0313-r. doi: 10.1094/mpmi-10-14-0313-r
[49]	LIU X, ZHU X L, WEI X N, LU C G, SHEN F D, ZHANG X W, ZHANG Z Y. The wheat LLM-domain-containing transcription factor TaGATA1 positively modulates host immune response to Rhizoctonia cerealis. Journal of Experimental Botany, 2019, 71(1): 344-355. doi: 10.1093/jxb/erz409. doi: 10.1093/jxb/erz409
[50]	ZHANG H, ZHANG Q, ZHAI H, GAO S P, YANG L, WANG Z, XU Y T, HUO J X, REN Z T, ZHAO N, WANG X F, LI J G, LIU Q C, HE S Z. IbBBX 24 promotes the jasmonic acid pathway and enhances Fusarium wilt resistance in sweet potato. The Plant Cell, 2020, 32(4): 1102-1123. doi: 10.1105/tpc.19.00641. doi: 10.1105/tpc.19.00641

引物名称 Primer name	引物序列 Primer sequence (5' to 3')	目的 Purpose
MdMDH-qpcr-F	CGTGATTGGGTACTTGGAAC	荧光定量PCR qRT-PCR
MdMDH-qpcr-R	TGGCAAGTGACTGGGAATGA
MD05G1054300-qpcr-F	CCGTACGGCAGCTGTAGAT
MD05G1054300-qpcr-R	GGAACTTCTCGAGGATTCGCA
MD05G1054400-qpcr-F	CGGAGGGGAAATCAAAGGCT
MD05G1054400-qpcr-R	TTGTACGGGCACCGACTAAC
MD05G1054500-qpcr-F	CCCATTTCGCCCTGTTCTGA
MD05G1054500-qpcr-R	AACACGCACCTACATCACCA
MD05G1048600-qpcr-F	GCAGGCCCAAGGACACTATG
MD05G1048600-qpcr-R	GCTACAACATCGGAATCCTGTT
MD05G1048700-qpcr-F	ACTCAACCATACGAGGCTCC
MD05G1048700-qpcr-R	ATTGCATGGGCAGCATGTTC
MD00G1039900-qpcr-F	GCTGCCCGATGTAAAACCAAA
MD00G1039900-qpcr-R	CCCACACCTTGTTCCTGGTT
MD00G1066800-qpcr-F	CAGAGTGGCAATGGTGGCTA
MD00G1066800-qpcr-R	TGCAAGTCGGTGGAGACTTC
MD05G1054300-attb-F	GGGGACAAGTTTGTACAAAAAAGCAGGCTGCatggagaacatcgcggcg	表达载体构建 Plasmid constructure of PK7-203
MD05G1054300-attb-R	GGGGACCACTTTGTACAAGAAAGCTGGGTCttatttccaaggaagcttaaacgtgt	表达载体构建 Plasmid constructure of PK7-203

染色体 Chromosome	显著关联SNP位置 Leading SNP position (bp)	基因型 Genotype	位置类型 Position type	-log10（P）值 -log10 (P)
Chr5	9339654	[T/C]	基因间区 Intergenic region	4.05E-08
Chr5	9357320	[T/C]	基因间区 Intergenic region	3.86E-08
Chr5	8263898	[T/C]	基因间区 Intergenic region	3.37E-08
Chr0	6821757	[A/G]	内含子 Intron	2.14E-08
Chr0	12882138	[T/C]	内含子 Intron	1.40E-08
Chr5	9336893	[C/G]	基因间区 Intergenic region	1.21E-08

SNP位置 SNP position (bp)	候选基因 Candicate gene	同源拟南芥 Homolog Arabidopsis	距离 Distance (bp)	基因功能 Gene function
9339654	MD05G1054300	AT2G43850	204058	整合素连接蛋白激酶 Integrin-linked protein kinase
9339654	MD05G1054400	AT4G38890	1049	FMN连锁氧化还原酶类超家族蛋白 FMN linked oxidoreductase superfamily protein
9357320	MD05G1054400	AT4G38890	3882	FMN连锁氧化还原酶类超家族蛋白 FMN linked oxidoreductase superfamily protein
9357320	MD05G1054500	AT4G38900	7806	碱性亮氨酸拉链（bZIP）转录因子家族蛋白 Basic leucine zipper (bZIP) transcription factor family protein
8263898	MD05G1048600	AT3G21175	12639	植物特异性GATA型转录因子的一个新家族成员 GATA-type transcription factor
8263898	MD05G1048700	AT1G21280	14343	多蛋白/逆转录转座子Multiprotein/retrotransposon
6821757	MD00G1039900	AT4G38960		B-box型锌指家族蛋白 B-box zinc finger family protein
12882138	MD00G1066800	AT4G34730		核糖体结合因子A家族蛋白 Ribosome-binding factor A family protein
9336893	MD05G1054300	AT2G43850	201297	整合素连接蛋白激酶 Integrin-linked protein kinase
	MD05G1054400	AT4G38890	3810	FMN连锁氧化还原酶类超家族蛋白 FMN-linked oxidoreductase superfamily protein