褪黑素预处理对桃耐涝性的调控效应

doi:10.3969/j.issn.1004-1524.2022.09.10

浙江农业学报 ›› 2022, Vol. 34 ›› Issue (9): 1911-1924.DOI: 10.3969/j.issn.1004-1524.2022.09.10

褪黑素预处理对桃耐涝性的调控效应

古咸彬(), 陆玲鸿, 宋根华, 肖金平, 张慧琴()

浙江省农业科学院园艺研究所,浙江杭州 310021

收稿日期:2021-11-24 出版日期:2022-09-25 发布日期:2022-09-30
通讯作者: 张慧琴
作者简介:*张慧琴,E-mail: zhqzaas@163.com
古咸彬(1988—),男,重庆江津人,博士,助理研究员,主要从事果树抗逆育种与分子生物学研究。E-mail: xbgu_8810@163.com
基金资助:
浙江省自然科学基金(LQ19C150003);国家自然科学基金(32102316);浙江省农业(果品)新品种选育重大科技专项(2021C02066-4);国家现代农业产业技术体系(CARS-30-Z-06)

Regulation effect of melatonin pretreatment on waterlogging tolerance in peach seedling

GU Xianbin(), LU Linghong, SONG Genhua, XIAO Jinping, ZHANG Huiqin()

Institute of Horticulture, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China

Received:2021-11-24 Online:2022-09-25 Published:2022-09-30
Contact: ZHANG Huiqin

摘要/Abstract

摘要：

为探究褪黑素对桃耐涝性的调控效应,用200 μmol·L^-1褪黑素对桃苗进行预处理,再经涝害处理,通过生理指标与转录组分析褪黑素和涝害对桃的影响。结果表明,涝害胁迫下外源褪黑素能保护细胞膜和根系。褪黑素能激活微管形态建成、氧化还原酶活性和蛋白结合相关的基因,巩固根系骨架,激活抗氧化酶活性,提高抗逆性。涝害诱导ERF转录因子家族成员大量表达,通过HRPE和GCC-box顺式作用元件激活下游ADH、ALD、GAPD等糖酵解途径关键酶基因的表达,为低氧胁迫下的植株供能。ERF Ⅶ成员Prupe.8G264900可能是桃响应涝害的关键基因。

关键词: 桃, 褪黑素, 涝害, 转录组

Abstract:

In order to explore the regulation effect of melatonin on waterlogging tolerance in peach, peach seedlings were pretreated with 200 μmol·L^-1 melatonin and then subjected to waterlogging treatment. Physiological indexes and transcriptome sequencing were used to study the effects of melatonin on peach under waterlogging. The results showed that exogenous melatonin slowed chlorophyll degradation and protected cell membranes and roots of peach seedling. Melatonin activated microtubule morphogenesis, REDOX enzyme activity and protein-binding related genes, thus consolidated root skeleton, activated antioxidant enzyme activity and improved stress tolerance. Waterlogging induced abundant expression of ERF transcription factor family members to regulate stress response. All the VII group members of ERF family were significant differential expressed in the waterlogging response group, which could be the key member in hypoxia stress response reaction. In addition, ERF transcription factor activated the activity of key enzymes genes as ADH, ALD, GAPD in glycolysis pathway through HRPE or GCC-box cis-acting elements, which provided energy for plants under hypoxic stress. Prupe.8G264900, a member of ERF Ⅶ, might be a key gene in response to waterlogging in peach.

Key words: peach, melatonin, waterlogging, transcriptome

中图分类号:

S662.1

古咸彬, 陆玲鸿, 宋根华, 肖金平, 张慧琴. 褪黑素预处理对桃耐涝性的调控效应[J]. 浙江农业学报, 2022, 34(9): 1911-1924.

GU Xianbin, LU Linghong, SONG Genhua, XIAO Jinping, ZHANG Huiqin. Regulation effect of melatonin pretreatment on waterlogging tolerance in peach seedling[J]. Acta Agriculturae Zhejiangensis, 2022, 34(9): 1911-1924.

图/表 12

表1 荧光定量PCR引物

Table 1 Primers for qRT-PCR

基因序列号 Gene ID	基因名称 Gene name	上游引物 Forward primer (5'→3')	下游引物 Reverse primer (5'→3')	产物长度 Products lenth/bp
Prupe.1G234000	PpGAPDH	CTGCTTCCTTCAACATCAT	TGCCTTCTTCTCAATCCT	158
Prupe.1G305900	PpRAV	TTCAGATGGTTCGGCTATT	TCCTTGGCTTCTTGCTAA	183
Prupe.2G258200	PpERD15	GAGGAAGAGGAGAAGGATT	CGGACTCACTTTCATCAC	138
Prupe.2G272300	PpERF1	CGGATGCTTCAACAACAA	TACTGGTCTTCTGCCTTG	111
Prupe.3G236200	PpLBD41	GGATGAGTTGTAATGGATG	CCTCGTATAGCAATGATC	213
Prupe.6G230600	PpWRKY7	CAGGTGTCTTCAGTTGGA	AGAGTAATCATCAGGTGGAA	207
Prupe.8G018100	PpADH1	GGAGTGATGCTGAGTGAT	CAGACCTGTGGAGATACC	183
Prupe.8G264900	PpERF071	CCTCCAATTCCAATTCCAA	GCAATCGCATCCTTATTCA	226
Prupe.6G163400	PpACTIN	TCTTCCAACCATCACTCAT	GCCACAACCTTAATCTTCAT	211

图1 淹水处理后桃生理指标的变化利用SPSS Statistics 17.0软件进行显著性差异分析,One-way ANOVA用于显著性检验,Duncan法进行多重比较。柱上无相同小写字母表示差异显著(P<0.05)。

Fig.1 Physiological indexes changes of peach after waterlogging treatment SPSS Statistics 17.0 software was used for significant difference analysis, one-way ANOVA was used for significance test, Duncan method was used for multiple comparison. Data on the bars marked without the same lowercase letter indicated significant differences at P<0.05.

图2 组间差异表达基因的统计分析

Fig.2 Number statistics of differentially expressed genes in groups

图3 褪黑素诱导富集的GO通路 A,GO富集分析;B,差异基因的表达热图。

Fig.3 Enrichment of the GO items induced by melatonin A, GO enrichment analysis; B, Heat map of differential gene expression.

图4 共同差异表达基因的GO富集分析与表达热图分析 A,组间比较Venn图;B,GO富集分析;C,差异基因的表达热图。

Fig.4 GO enrich and expression analysis of common differentially expressed genes A, Venn diagram between groups; B, GO enrichment analysis; C, Heat map of different expression gene.

表2 GO显著富集通路中差异表达的转录因子

Table 2 Different expressed transcription factors in the most enrich GO pathway

转录因子家族 TF family	数量 Number	转录因子家族 TF family	数量 Number
AP2/ERF	34	HD-ZIP	3
WRKY	23	NF-YA	3
bZIP	15	ARR-B	2
HSF	12	BES1	2
C2C2-GATA	4	NF-X1	1
MADS	4	SBP	1
AT-hook	3	Trihelix	1

图5 桃和拟南芥ERF家族进化树分析

Fig.5 Phylogenetic tree analysis of ERF family in peach and Arabidopsis thaliana

表3 桃ERF转录因子家族成员差异表达基因分布

Table 3 Different expressed ERFs in each subfamily group

ERF家族 ERF family	数量 Number	百分比 Percentage/%	ERF家族 ERF family	数量 Number	百分比 Percentage/%
Ⅰ	1/6	16.7	Ⅶ	3/3	100
Ⅱ	1/8	12.5	Ⅷ	3/10	30
Ⅲ	5/22	22.7	Ⅸ	12/20	60
Ⅳ	2/7	28.6	Ⅹ	2/6	33.3
Ⅴ	0/12	0	Ⅺ-L	0/2	0
Ⅵ	0/6	0

图6 差异表达的ERF成员的FPKM统计

Fig.6 FPKM number of different expressed ERF family members

图7 KEGG富集分析及ERF靶基因预测 A,KEGG富集气泡图;B,糖酵解途径与合成酶基因的表达热图;C,ERF靶基因预测。

Fig.7 KEGG pathway enrichment analysis and ERF transcription factor target gene prediction A, KEGG enrichment analysis; B, Expression heat map of glycolysis pathway genes; C, ERF target gene prediction.

图8 差异表达基因的荧光定量表达验证

Fig.8 qRT-PCR validation of relative expression levels of differential expressed genes

图9 桃响应涝害调控网络图

Fig.9 Hypothesized regulatory mechanism for melatonin in the waterlogging response pathway

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褪黑素预处理对桃耐涝性的调控效应

Regulation effect of melatonin pretreatment on waterlogging tolerance in peach seedling

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