摘要
【目的】芍药在生长发育过程中容易遭受病原菌侵染,感染病害,由细极链格孢(Alternariatenuissima)引起的红斑病是其主要叶部病害,严重影响品质和产量,目前对芍药红斑病的抗病机理尚未清晰。本研究运用生理学和转录组学手段,探究芍药响应细极链格孢侵染的生理变化及分子途径。【方法】以芍药‘大富贵’为试验材料,取A.tenuissima接种后12、24和96 h的芍药叶片,测定其相关生理指标并进行转录组测序分析,以未接种叶片为对照。【结果】病原菌接种后,芍药叶片的超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)和苯丙氨酸解氨酶(PAL)活性增强,可溶性糖与可溶性蛋白以及丙二醛(MDA)含量升高,脯氨酸含量降低。在A.tenuissima侵染芍药12、24和96 h后,芍药分别有5045、5961和2748个差异表达基因上调,有4284、5665和3536个差异表达基因下调。GO富集分析发现差异基因主要富集到部分生物合成和代谢过程、光合作用、信号转导和节律相关条目中。KEGG富集分析显示差异基因主要富集到碳代谢、氨基酸的生物合成、植物-病原菌互作、植物激素信号转导、MAPK信号等通路中。3条抗病通路(植物-病原菌互作、MAPK信号和植物激素信号转导)中共同富集到53个差异基因,这些差异基因中包括1个MPK6、2个PR1、4个MKK4/5和46个BAK1。随机选取9个芍药响应细极链格孢侵染的差异表达基因进行qRT-PCR分析,基因表达规律与转录组测序结果一致,证实RNA-seq的准确性。AP2/ERF-ERF、WRKY、bHLH和MYB-related转录因子家族是芍药响应A.tenuissima侵染的关键转录因子家族。【结论】A.tenuissima侵染芍药后,芍药通过提高SOD、POD、CAT和PAL的活性提升抗氧化能力,清除病害胁迫所产生的大量活性氧,通过增加可溶性糖与可溶性蛋白的含量、降低脯氨酸的含量进行渗透调节保持细胞水分。BAK1、PR1、MKK4/5和MPK6在芍药响应A.tenuissima侵染过程中起到重要作用,推测PlERF20、PlERF1b、PlWRKY41、PlMYC4和PlMYB62是芍药响应A.tenuissima侵染的抗病相关转录因子。
【Objective】Paeonia lactiflora(P.lactiflora)is prone to pathogen infection during its growth and development.Leaf red spot disease caused by the Alternaria tenuissima(A.tenuissima)seriously affects both the quality and yield of the plant.However,the disease resistance mechanism of P.lactiflora is not clear at present.This study aimed to explore the physiological changes and molecular response pathways of P.lactiflora in response to the infection of A.tenuissima by using physiological and transcriptomic approaches.【Method】Taking P.lactiflora Dafugui as the experimental material,the leaves of them at 12,24 and 96 h after A.tenuissima infection were taken respectively to determine the relevant physiological indexes and perform transcriptome sequencing analysis,used uninoculated leaves as the control.【Result】After pathogen inoculation,the activities of SOD,POD,CAT and PAL of the leaves of P.lactiflora increased,the content of soluble sugar,soluble protein and MDA increased,but the content of proline decreased.The differentially expressed genes(DEGs)in each stage were screened after 12,24 and 96 h of A.tenuissima infection.There were 5045,5961 and 2748 DEGs up-regulated,and 4284,5665 and 3536 DEGs were down-regulated in each stage.GO enrichment analysis showed that these DEGs were mainly enriched in some biosynthesis and metabolic processes,photosynthesis-related,signal transduction-related and rhythm-related items.KEGG enrichment analysis showed that these DEGs were mainly enriched in carbon metabolism,amino acid biosynthesis,plant-pathogen interaction,plant hormone signal transduction,MAPK signaling,and other pathways.A total of 53 DEGs were enriched in three disease resistance pathways(plant-pathogen interaction,MAPK signaling and plant hormone signal transduction pathways).These DEGs included 1 MPK6,2 PR1,4 MKK4/5,and 46 BAK1.Nine differentially expressed genes of P.lactiflora in response to A.tenuissima infection were selected for qRT-PCR analysis.The gene expression pattern was consistent with the transcriptome sequencing results,which confirmed the accuracy of RNA-seq.The transcription factor families analysis showed that the AP2/ERF-ERF,WRKY,bHLH and MYB-related transcription factor families were the key transcription factor families of P.lactiflora in response to A.tenuissima.【Conclusion】After A.tenuissima infected,the antioxidant capacity of P.lactiflora was increased by increasing the activities of SOD,POD,CAT,and PAL,to remove the large amount of reactive oxygen species produced by disease stress,maintaining cell water by increasing the content of soluble sugars and soluble proteins and reducing the content of proline.The continuous increase of MDA content indicated that the cell membrane structure of plants was destroyed.The BAK1,PR1,MKK4/5 and MPK6 genes played important roles in response to A.tenuissima infection.It was preliminarily speculated that PlERF20,PlERF1b,PlWRKY41,PlMYC4,and PlMYB62 were disease resistance related transcription factor in response to A.tenuissima infection.
作者
蔚书涵
秦小杰
吴其超
李玲
臧德奎
马燕
YU ShuHan;QIN XiaoJie;WU QiChao;LI Ling;ZANG DeKui;MA Yan(College of Forestry,Shandong Agricultural University/Key Laboratory of State Forestry Administration for Silviculture of the Lower Yellow River,Taian 271018,Shandong)
出处
《中国农业科学》
CAS
CSCD
北大核心
2024年第15期3035-3052,共18页
Scientia Agricultura Sinica
基金
山东省自然科学基金(ZR2020MC162)
山东省重点研发计划(重大科技创新工程)(2021LZGC023)
山东省林业改革发展资金专项(鲁财预指[2021]1号)。
关键词
芍药
细极链格孢
转录组学
生理指标
抗病通路
Paeonia lactiflora Pall.
Alternaria tenuissima
transcriptomics
physiological index
resistance pathways
