Screening And Identification Of Sclerotiniose Response In Mulberry Small Secretory Peptides And Key Genes

The mulberry contains a variety of nutrients and bioactive substances,and has high nutritional and medicinal value,and is included in the "food and medicine" category.This thesis focuses on the comparative transcriptome analysis of different mulberry species and different pathogenic fungi,screening for small secretory peptides and candidate genes in response to mulberry sclerotiniose,using a virus-induced gene silencing system for gene function validation,and selecting the gene family in which the key candidate gene MaSWEET1a is located for bioinformatic analysis and expression profiling.The gene family of MaSWEET1a was selected for bioinformatics analysis and expression profiling to explore its potential role in the mulberry-mulberry sclerotiniose pathogen cupping interaction.The main findings are as follows:(1)The conditions of the virus-induced silencing system in mulberry were optimised for three inoculation methods,co-culture time,vacuum time,vacuum pressure,state of infested plants and Agrobacterium tumefaciens infestation concentration,and the fluorescent signal was used to confirm vector expression to reduce the workload of the later RT-qPCR screening of silenced plants,and the mulberry virus-induced gene silencing system was successfully optimised for mulberry gene function The system was successfully optimized to provide technical support for the functional validation of mulberry genes.(2)Using mulberry transcriptome data from public databases,1088 small secreted peptides were identified and annotated in mulberry,and a total of 2956 differential genes were screened by comparing the transcriptome data of resistant variety K1(K),healthy fruit of Morus atropurpurea variety Zhongshen1(L)and fruit infected with Ciboria shiraiana(S).52 candidate genes and small secreted peptides,named mycorrhizal response genes(SRGs),were screened by comparative transcriptome analysis of different mulberry species infected with different pathogenic fungi.(3)6 SRGs were selected for gene function validation,and three negative(MaSSP15,MaSWEET1a,MaMES17)and three positive(MaPPO,MaAOS,MaLOX1)regulators of resistance to mulberry sclerotiniose infection were identified using VIGS silencing of the selected 6 SRGs in mulberry and back-infection experiments with Morus alba.(4)Further bioinformatic analysis and expression profiling of the gene family in which MaSWEET1a is located was carried out.A total of 24 genes of SWEETs were identified in mulberry and classified into 4 categories.Most of the MaSWEETs were distributed in the plasma membrane,vesicle membrane and chloroplast-like vesicle membrane.They were distributed on all but chromosomes 1 and 13.Multiple cis-acting elements responding to hormones(e.g.,jasmonic acid,abscisic acid,gibberellin,etc.)were identified by promoter region analysis,and genes from adjacent branches had similar gene structures.The expression patterns of SWEETs genes in different developmental branches in response to mulberry sclerotiniose(infection with Ciboria shiraiana)are not identical or even opposite,and further investigation is needed to investigate how the pathogen controls the expression of these genes.