The Physiological And Molecular Mechanism Of Sea Buckthorn(Hippophae Rhamnoides) In Response To Drought Stress And Rehydration

Sea buckthorn(Hippophae rhamnoides)belongs to hippophae of elaeagnaceae,which is widely distributed in semi-arid regions of China.With the global climate changing,the temperature of Asia is rising obviously and the precipitation is decreasing.The sea buckthorn is facing the more severe drought climate challenge.There are a series of mechanisms,which response to drought stress,having been formed in the long term evolution of plants.At present,although the physiological and biochemical of sea buckthorn responsing to drought stress and proteomics have been studied,the research which illuminate the molecular mechanism of sea buckthorn responses to drought stress has not been carried out in the level of gene expression and regulation.Based on the above research,this study chosen two cultivars of sea buckthorn(H.rhamnoides L.subsp.Sinensis,‘Fengning',FN and H.rhamnoides L.subsp.Mongolia,‘Xianyang',XY)which difference in drought resistance to simulate soil drought and rehydration test of natural environment.In this process,the plant water relations and photosynthetic physiological characteristics were measured in different drought degree and the difference and commonness that response to drought were compared.The leaves of sea buckthorn under three conditions(normal water,severe drought stress and rehydration)were used for the transcriptome sequencing and analysis of data.The signal transduction and gene regulation mechanism of sea buckthorn in response to drought stress were revealed by the gene expression level.Western blot was used to verify the binding capacity of H3K9 ac antibody and the histone of sea buckthorn,and antibody was used for subsequent Ch IP enrichment experiments.The enriched DNA fragments were high throughput sequenced and date were used to map the H3K9 acetylation of sea buckthorn.The regulatory genes of H3K9 acetylation were also identified.The main results are showed as follows:(1)Leaves,experiencing drought treatment,initially shrunk at 22 nd(soil moisture voleme content 19.40%)for XY and 33 nd(11.66%)for the FN.In addition,with the increasing drought stress,the leaf water potential of both species decreased while the moistureabsorption ability increased.The maximum net photosynthetic rate,net photosynthetic rate,transpiration rate,stomatal conductance,light saturation point,and apparent quantum yield decreased with increasing drought stress;whereas,the light compensation point increased.Water efficiency also rose after falling.The dark respiration rate of FN treatment group decreased first and then recovered.But XY treatment group unchanged first and then increased.After rehydration,the physiological indexes appeared in different degrees of recovery.Under the drought stress,the morphological characteristics,water potential and photosynthetic physiology of FN and XY obvious changed which reflected the changes of sea buckthorn in natural arid environment.Although sea buckthorn is a hardy plant,its drought tolerance is limited.Therefore,when extreme drought occurred in the process of cultivating a plantation,the leaves shrunk and need timely irrigation to avoid unrecoverable damage with FN being more suitable than XY as a breeding parent.(2)Drought stress seriously affect the gene expression of sea buckthorn leaves.FN and XY were found in 4618 and 6100 differentially expressed genes(DEGs),respectively.After rehydration,the number of DEGs less quickly and there are seven and 26 respectively.This results showed that drought stress on the influence of DEGs and recovery ability between different cultivars.The analysis of GO and KEGG of DEGs found that the two cultivars of sea buckthorn have similar response mechanism in response to drought stress.There is a significant enrichment of photosynthetic system related functions,which explains the mechanism of significant change in net photosynthetic rate.Predicted and identified of 3170 new genes and 970 transcription factors improved and enriched the sea buckthorn genome information.The new genes were involved in the 101 metabolic pathways which metabolic synthesis pathway contains the most genes.Transcription factors were distributed among 71 gene family and the MYB family has the largest number of genes.Based on gene co-expression network building and public drought database screening,there are 194 core drought stress response genes were filtered.The analysis illustrates the drought tolerance mechanism which include mainly ABA dependent signaling pathway in signal transmission way and flavonoids biosynthesis pathway playing roles in oxygen free radical removal.Inaddition,the new relationship between the two pathway was found and the gene expression regulation mechanism was enriched.The protein interaction of transcription factors were used to supplement the drought stress mechanism.(3)The strong binding ability of H3K9 ac antibody and compound of protein and DNA were proved by western blot.The enrichment and high throughpur sequencing of the DNA fragment prove that antibodies can be used to research the types of histone modification.FN and XY in 2.2x107 amd 2.5x107 original reads by sequencing.The unique mapped reads is widely distributed in the sea buckthorn genome and there is an obvious enrichement of both ends of the genebody.FN and XY predicted 1011 amd 8670 peaks respectively,and the first H3K9 ac modified genetic map was drawn and the genes were indentified modifing by H3K9 ac.FN contains 332 specific loci,XY contains 4975 specific loci and 195 loci exist both at the two cultivars.The gene function of specific site area includes multiple biological processes.The ABA dependent signaling pathway and flavonoids biosynthesis that responses to drought stress were regulated by H3K9 ac.This information provide a new epigenetic insights for the difference between cultivars and lay a foundation for the further study of the regulation of gene expression and the expression of gene expression in response to envitonment stress.