摘要
【目的】灰斑病是枇杷生产中对产量和品质影响最为严重的病害之一,探讨枇杷抵御灰斑病菌侵染的分子机制,并筛选灰斑病菌侵染枇杷叶片时的应答基因,为抗性育种提供基因基础。【方法】利用田间调查及人工离体叶片接种的方法,对收集保存的52份枇杷种质的灰斑病抗性进行评价;以接种灰斑病菌后33、36和48 h的枇杷幼嫩叶片为试材,利用转录组测序技术对枇杷抗灰斑病相关基因和代谢途径进行分析。【结果】在52份枇杷种质中,田间调查和人工离体接种结果一致的有11份。接种后33、36和48 h在抗感品种中共检测到684个差异表达基因,其中上调基因为585个,下调基因为99个,一共有25个(12个上调和13下调)差异表达基因仅在抗病品种白梨的3个处理中差异表达。GO富集分析结果显示,枇杷叶片响应灰斑病侵染的差异表达基因,主要集中在对外源性刺激的反应、植物型次生细胞壁生物发生和光合作用光反应等过程中。KEGG富集分析结果显示,高抗与高感种质的差异表达基因多数富集在次生代谢产物的生物合成、MAPK信号通路-植物以及植物-病原体相互作用等通路。WRKY40-like等7个基因的RT-qPCR验证结果与RNA-seq数据一致,且这些基因在接种后33、36和48 h均能响应灰斑病菌的侵染。【结论】从转录组水平上分析了枇杷响应灰斑病菌侵染过程中的一些关键基因,其中高抗种质的差异表达基因中存在AP2/ERF类转录因子和3个抗病基因。本研究结果将有助于优化枇杷抗灰斑病菌的调控机制,为深入探究枇杷与灰斑病病原菌互作分子机制提供参考依据。
【Objective】Gray spot disease is one of the most serious diseases affecting the yield and qual-ity of loquat in the production.This study investigates the molecular mechanisms underlying loquat re-sistance to Pestalotiopsis infection and identifies the responsive genes in loquat leaves during pathogen invasion,aiming to lay the genetic foundation for resistance breeding.It is generally believed that the causative agent of loquat gray spot disease belongs to the deuteromycete,melanospora,and the pestalo-tiopsis,which can infect several organs of loquat,and when it infects the leaves,they become rigid and fall easily;when it infects the flowers,fruits and root necks,it causes them to rot,which has a great im-pact on the growth and development of the tree and leads to a serious decrease in the yield and quality of loquat.Currently,loquat gray spot is mainly defended and controlled using chemical agents,but it tends to lead to increased resistance of the pathogen and pollution of the environment.Therefore,im-proving the disease resistance of loquat germplasm is the most economical and effective way to solve this problem,and a good assessment of the disease resistance of different loquat germplasm is a prereq-uisite for breeding disease-resistant varieties,which is also important for the study on the mechanism of loquat response to infestation by the gray spot pathogen and breeding for disease resistance.At present,due to the lack of disease-resistant germplasm and its genetic information,there are few studies on the mechanism of interactions between loquat and pestalotiopsis,especially on the pathogenicity of patho-genic bacteria and the mechanism of plant disease resistance,and the breeding of disease-resistant germ-plasm is progressing slowly.In this study,loquat gray spot disease was taken as the research object to explore the molecular mechanism of loquat resistance to the infection of gray spot pathogen,and to screen the relevant response genes when gray spot pathogen damaged loquat leaves,so as to provide the genetic basis for resistance breeding.【Methods】The resistance of 52 loquat germplasm resources col-lected and preserved to Cercospora cinerea was evaluated by using software and visual methods,com-bined with the method of artificial in vitro leaf inoculation.In addition,transcriptome sequencing tech-nology was used to analyze the changes of gene expression levels in leaves of highly susceptible and highly resistant germplasm resources after inoculation,and to screen the main disease-resistance-related genes,so as to find out the disease resistance factors of loquat germplasm,in order to reveal the disease resistance mechanism of loquat gray spot disease,and provide some guidance and theoretical basis for the breeding of loquat gray spot disease resistance.【Results】Among the 52 loquat germplasm resourc-es,eleven were consistent with the results of field investigation and artificial inoculation in vitro,includ-ing three high-resistance germplasms(Baili,Eriobotrya fragrans and E.serrate);Four disease-resistant germplasms(E.fulvicoma Chun&Liao,E.deflexa f.koshunensis,Mogi,Shiromogi);Two susceptible germplasms(Dahongpao and E.kwangsiensis);And two highly susceptible germplasms(Guangrong-ben and E.cavaleriei.).By analyzing the transcriptome of the leaves of the inoculation treatments and the control of the high resistance germplasm'Baili'and the high sensitivity germplasm Guangrongben with loquat leaf gray spot disease,a total of 8664 DEGs were detected in the comparison of BL-33h and BL-33ck of Baili cultivar,of which 4049 were up-regulated and 4615 were down regulated.In the com-parison of subsequent time points,both the number of up-regulated and down regulated DEGs showed a downward trend,while Guangrongben variety was just the opposite.In the comparison of GRB-48h and GRB-48ck,there were a total of 7483 DEGs,3098 of which were up-regulated and 4385 were down regulated.It is speculated that the difference in the number of DEGs between the two varieties at differ-ent time points may be related to their different resistance to gray spot disease.The analysis found that a total of 684(585 up-regulated and 99 down regulated)DEGs responded in Baili and Guangrongben,and a total of 25(12 up-regulated and 13 down regulated)DEGs responded only in Baili.GO and KEGG analysis were performed on these co-responsive genes and special responsive genes,and their transcription factors and disease resistance genes were identified.64 transcription factors and 59 disease resistance genes were identified in the co-responsive genes,and one transcription factor and three dis-ease resistance genes were identified in the special responsive genes.【Conclusion】In this study,52 lo-quat germplasm resources were evaluated for resistance to Cercospora cinerea more accurately,and some key genes in response to Cercospora cinerea infection in loquat were analyzed at the transcrip-tome level.AP2/ERF transcription factors and three disease resistance genes were found in the differen-tially-expressed genes of high resistance germplasm.The results could help to enrich the regulation mechanism of resistance to Cercospora cinerea in loquat.The differences in resistance related pathways between resistant and susceptible germplasm resources were helpful to reveal the reasons for the differ-ences in resistance.The key metabolic pathways and resistance-related genes in response to Cercospora cinerea infection in Loquat germplasm with high resistance were identified.The molecular mechanism of pathogen interaction provides a reference for the theory.
作者
陈依丽
李文祥
陈依莹
彭泽
杨向晖
CHEN Yili;LIWenxiang;CHEN Yiying;PENG Ze;YANG Xianghui(Baise Modern Agricultural Technology Research and Promotion Center,Baise 533612,Guangxi,China;College of Horticulture,South China Agricultural University,Guangzhou 510642,Guangdong,China;College of Horticulture,Nanjing Agricultural University,Nanjing 210000,Jiangsu,China;The College of Mathematics Sciences,Harbin Engineering University,Harbin 150001,Heilongjiang,China;Utilization of Horticultural Crop Resources in South China,Ministry of Agriculture and Rural Affairs,Guangzhou 510642,Guangdong,China)
出处
《果树学报》
北大核心
2025年第12期2940-2960,共21页
Journal of Fruit Science
基金
广东省重点领域研发计划项目(2022B0202070002)
广州市科技计划项目(202206010147)。
关键词
枇杷属
灰斑病
抗病评价
转录组
差异表达基因
Eriobotrya
Gray spot disease
Disease resistance evaluation
Transcriptome
Differentially expressed genes
