Screening Of Key Target Genes Under High Temperature Stress In Lasiodiplodia Theobroma

Our research group found Lasiodiplodia theobromae can induce tea leaf spot on tea plantation in Guizhou Province.L.theobromae is a high temperature type pathogen,but the host–pathogen interactions between L.theobromae and tea under high temperature stress has not been reported.In this paper,various analyses were used to find the key target genes about temperature,which will contribute to studying the host–pathogen interactions and on disease resistance mechanisms.In order to study the biological characteristics of pathogen,the tea leaves were inoculated by L.theobromae,and then the light brown lesions gradually enlarged into dark brown,irregular lesions.L.theobromae was transferred into the medium with different carbon sources,which found that the hyphae on the glucose medium grew best.L.theobromae was transferred into the medium with different nitrogen sources,and found that the hyphae on the leucine medium grew best and the abundance of mycelia in potassium nitrate was higher.In order to observe the morphology of hyphae,it was found by scanning electron microscope that the extracellular lipopolysaccharide secreted by hyphae in different media was different.In order to study the mycelium morphology,membrane and nucleus can be observed by diisopropanolamine and calcium fluorescent white staining.The effects on the ultrastructure of tea leaves during the infection of pathogen was observed by transmission electron microscope.When tea leaves were infected by pathogens,the morphological structure of palisade tissues and sponge tissues changed and the cell wall was incomplete,particle accumulation increased,and the intracellular edges were obvious.In order to screen out the key target genes,the transcriptome was used to study the gene expression of L.theobromae under the stress under different temperatures.1476 and 2415 of the differentially expressed m RNAs(DEm RNAs)were up-or downregulated.The Kyoto Encuclopedia of Gene and Genomes(KEGG)enrichment analysis of the DEm RNAs were enriched in the Ribosome biogenesis in eukaryotes pathway;lnc RNAs were used to analyze the gene expression of L.theobromae after infection of tea leaves.3403 and 1477 of the differentially expressed lnc RNAs(DElnc RNAs)significantly up-or down-regulated,which cis-regulated lnc RNAs and m RNAs were 176 and 146,respectively.For the term of defense response at the Gene Ontology(GO)aspect of biological process,Glycosyl hydrolase 9A1 was cis-regulated by the DElnc RNAs;circ RNA was used to study the gene expression of L.theobromae after infection of tea leaves.92 and 153 of the differentially expressed circ RNAs(DEcirc RNAs)were significantly up-or down-regulated.The DEcirc RNAs were mainly enriched in DNA binding in GO database.WRKY transcription factor 24 was mainly enriched in the pathway of plant-pathogen interaction in the KEGG enrichment analysis.In the mi RNA analyse,the differentially expressed mi RNAs were mainly enriched in transcription,DNA-templated,and oxidation-reduction process protein phosphorylation.For the term of defense response,the differentially expressed genes(DEGs)were mainly enriched in MAPK signaling pathway-yeast.2373 lnc RNAs and606 cir RNAs were identified,and PC-3p-3047034＿3 targeted transcription factor Hsf B3a in the ce RNA analysis.At the same time,the mi RNAs PC-5p-1745146＿4,PC-5p-1706779＿4,and PC-3p-2090095＿4 targeted the same m RNA.In the analysis of degradome data,mi RNA PC-3p-3047034＿3 targeted TEA000588.1.mi RNA GMAmir1520L＿L-2R-4＿1SS3TG targeted TEA029729.1 which was annotated as Hsp90-1,and PC-5p-2230964＿3 targeted TEA003992.1 which was annotated as heat shock cognate protein 80.Semi-quantitative and RT-qPCR methods were used to verify the DEGs.In this study,L.theobromae infected tea leaves under the stress of high temperatures and key genes were selected,which was helpful to study host–pathogen interactions and on disease resistance mechanisms.