Funational Study On Serendipita Indica Promote Sseed Germination And Protocorm Growth Of Orchids

Mycorrhiza is a symbiont formed between plant roots and fungi,which can be divided into ectomycorrhiza and endophytic mycorrhiza.Endophytic mycorrhizae include Arbuscular mycorrhizae(AM),Orchid mycorrhiza(OM)and Ericoid mycorrhizae(ERM).The Orchid mycorrhiza fungi mainly includes several groups of Sebacinales,Ceratobasidiaceae and Tulasnellaceae.The Serendipita indica belongs to Sebacinales,which is symbiotic with extensive plants.At present,little research has been done on the symbiotic relationship between S.indica and orchids.In this study,S.indica was used as the experimental strain to conduct symbiosis experiments with the seeds of a variety of orchids to study the function of symbiosis between S.indica and orchids.The function of S.indica in the growth of Orchidaceae was discussed in combination with the effect of promoting seed germination,the microscopic structure of the infected plants,the level of transcriptional regulation.The research results are as follows:(1)S.indica can promote seed germination to seedling stage of various orchids.First of all,through the symbiotic germination experiments with 13 species of orchids,it was concluded that S.indica could promote Dendrobium.officinale,D.nobile,D.chrysotoxum,D.wangliangii,Cymbidium bicolor and C.aloifolium seeds germinate and grow to the seedling stage.The symbiotic germination lasted 90 days:seed germination rate and seedling formation rate is D.officinale(84.6 ± 5.2 %,78.1 ±5.5 %),D.nobile(67.4 ± 11.5 %,61.7 ± 12.4 %),D.wangliangii(46.2 ± 1.9 %,36.8± 4.2 %),C.aloifolium(41.4 ± 6.4 %,32.4 ± 5.1 %),D.chrysotoxum(24.4 ± 8.2 %,23.2 ± 7.8 %),C.bicolor(8.6 ± 4.6 %,7.3 ± 4.1 %).(2)S.indica forms endophytic mycorrhiza in orchids.Through microscopic observation,it was found that S.indica existed in the protocorm cells of orchids in the form of mycelia and mycelia clusters.When the protocorm grew into the adult plant,S.indica was still found in the root.The hyphae of S.indica was found to attach to the root surface,penetrate the root cortex cells and reach the cortex cells of host orchid(D.officinale).The chlamydospore of S.indica were found on the root surface and root cells of D.officinale.(3)S.indica had similar effects on seed germination of D.officinale with endophytic fungi LQ and JM.Then,two endophytic fungi,LQ and JM,which can promote seed germination of D.officinale were selected and compared with S.indica.It was found that the germination promotion effect of S.indica and the endophytic fungi of Orchidaceae were similar.After 90 days of symbiotic sowing,the germination rate,protocorm formation rate and seedling formation rate of the three strains of fungi under the treatment were::S.indica(93.4 ± 4.8 %,88.8 ± 5.7 %),LQ(72.4 ± 8.3 %,64.1 ± 5.8 %),and JM(66.6± 10.8 %,45.3 ± 9.4 %).(4)The level of Extracellular ATP in D.officinale is significantly up-regulated during the symbiosis between S.indica with D.officinale.Then,Extracellular ATP(e ATP)of D.officinale was detected at the 1 dpi(days post inoculation),2dpi,3dpi and 8dpi of symbiosis between S.indica and protocorm.The results showed that the e ATP level of D.officinale peaked on 2dpi and was low from the 3dpi to 8dpi.The expression levels of CPK28 and RBOHD,the downstream genes related to ATP signal transduction,were detected,and the expression levels of both marker genes were upregulated 1-3 folds under the treatment of S.indica.With the change of EATP level,the quantity of fungal colonization in host plants increased significantly from the second day of colonization and leveled off to the eighth day of colonization,but it was still at a high level.Extracellular ATP in plants usually plays a role in signal transduction related to defense.We hypothesized that the signaling pathway of e ATP may be involved in regulating the symbiosis between D.officinale with S.indica.(5)Transcriptome analysis of different stages of symbiosis between S.indica with D.officinale.Finally,transcriptome sequencing was used to obtain the differentially expressed genes in the early(3dpi),middle(9dpi),and stable(18dpi)stages of the symbiosis,and46 differentially expressed genes in three different symbiosis stages were identified.These genes will be used as the target genes related to the symbiosis process,and the preliminary gene annotation analysis will open the direction for the future research on the molecular mechanism of the symbiosis relationship between D.officinale and S.indica.