Study On The Auxin Regulation Of The Initial Culm Gall Formation Of Zizania Latifolia

Zizania latlfolia also known as Jiaogua,Jiaosun and Gaogua et al,is a traditional high-quality aquatic vegetables.Z.latlfolia is formed after the infection of Ustilago esculenta,which prevents its plants from heading and flowering,and stimulates the culm hypertrophy.The initial triggering stage and the later swelling stage are two different processes in culm gall formation of Z.latlfolia.Previous studies have suggested that auxin(IAA)and zeatin(ZT)played an important role in the expansion of swollen culm,but how the culm gall started to swell and the pattern of hormone changes in this process was not yet clear.Therefore,this paper was focused on the study of the initial triggering stage of Z.latifolia formation.In this study,U.esculenta "UeYD-3" was isolation form Z.latlfolia"Z1YD-3",and the whole genome sequence of the strain was assembled by the new generation sequencing technique.Based on the genome of the first version of Z latlfolia(INSDC:ASSH00000000.1)and the U.esculenta "UeYD-3"(INSDC:JAAKGJ010000000),the transcriptome analysis of interaction between Z latlfolia and U.esculenta in the growth points in seedling stage(5 leaves stage)and early swelling stage(8 leaves stage)of Z.latlfolia were analyzed,and the differences of hormone content were also compared between them.Furthermore,the key hormones and related genes regulating the formation of Z.latifolia were identified by qRT-PCR,and RNAi interference vectors were constructed.The purpose of this paper was to find out the role of of hormone in the process of initial swelling of Z.latifolia,and to provide the theoretical basis for regulating clum gall formation and guiding the production of Z.latifolia.The results are as follows:1.Using the sequencing technology combining the second-generation Illumina platform and the third-generation PacBio platform,the whole genome of the U.esculenta"UeYD-3" was assembled and the gene function annotation and comparative genomic were analyzed.It was found that the genome of the U.esculenta "UeYD-3" was assembled into 34 Scaffolds with a size of 26,430,365bp,N50 of 1,029,683bp,N90 of 465,370bp,and GC content of 53.12%.A total of 7091 genes were encoded in the genome,with a gene annotation rate of 95.5%.Analysis of common genes and unique genes between U.esculenta.YD-3 and maize U.maydis.521,maize S.reilianum.SRS1.H2-8 and brome U.bromivora.UB2112.The results showed that there are 4,525 identical genes in the four strains,With a length of 2,950,982bp;9,870 non-redundant genes,with a length of 5,887,468bp;and 932 unique genes of U.esculenta.Analysis of phylogenetic tree show that U.esculenta.YD-3 has high homology with the maize U.maydis.521 strain and the maize S.reilianum.SRS1.H2-8 strain,and the genetic distance with the brome U.bromivora.UB2112 strain was far.2.Analysis of mixed transcriptomes between Z latlfolia and U.esculenta at the seedling stage and initial swelling stage of Z latlfolia,and the average output data of each sample is 15.25Gb.The average comparison rate of the samples against the U.esculenta"UeYD-3",genome was 4.76%,and a total of 6,330 genes were obtained,and there were 243 differential genes,including 99 up-regulated genes and 135 down-regulated genes.The average comparison rate of the samples against the Z latlfolia genome was 84.03%,a total of 40,856 genes were obtained,and 4,936 differential genes were included,including 2,164 up-regulated genes and 2,772 down-regulated genes.3.Using the genome of strain U.esculenta "UeYD-3" as the reference genome,the biological pathway category in GO function mainly focuses on genes related to catalytic activity;The KEGG enriched metabolic pathway mainly focuses on the biosynthesis of antibiotics.Based on the genome of Z latlfolia,the biological pathway category in GO function mainly focuses on genes related to metabolic process and cell process;GO functional classification mainly focuses on cell-related genes in the cell component category,including 879 up-regulated genes and 543 down-regulated genes;1345 binding-related genes and 1471 gene-related genes in molecular function categories;The KEGG enriched metabolic pathways mainly focused on plant hormone signaling transduction,with 185 differentially expressed genes.4.The enlargement of the fleshy stems of Z.latifolia is closely related to the changes of hormone levels.In this study,it was found that there were significant differences in gene expression levels related to plant hormones between the seedling stage and initial triggering expansion of Z latlfolia.There were 95 differentially expressed genes related to hormone synthesis,degradation and signal transduction,including 58 up-regulated genes and 37 down-regulated genes.Further analysis showed that the expression of Zlat 10038559,a coding gene of YUCCA(a flavin monooxygenase,catalyzes a key step in tryptophan-dependent auxin biosynthesis),was significantly increased.Moreover,Local BlastP analysis of YUCCA gene sequences from Oryza sativa and Arabidopsis thaliana was conducted in the genome of Z.latlfolia,and a total of 13 YUCCA family genes were obtained,the amino acid sequence of Zlat 10038559 has the highest homology and closest genetic relationship with rice OsYUCCA9,and named ZlYUCCA9.In addition,IAA content of the Z.latlfolia at the initial triggering of culm swelling was significantly increased,and ZT,GA1,GA3,ABA,JA,SA were all unresponsive or down-regulated.All these results indicated that auxin plays an important role in the initial of Z.Latifolia formation.5.The construction of the ZlYUCCA9 RNAi vector by the special chemical induction expression vector pX6-GFP.It could be used to further study the function of YUCCA genes in initial of culm gall formation of Z latlfolia.