Transcriptomics-based Physiological Mechanism Of Barley Resistance To Bipolaris Sorokiniana And Screening Of Disease Resistance Genes

Barley(Hordeum vulgare L.)is the fourth cereal crop in the world.It not only has short growth period and strong adaptability,but also possesses the cold tolerance,drought tolerance,poor tolerance and salt and alkali resistance.Which is also widely distributed all over the world,especially for most areas of China.Spot blotch is a fungal disease caused by Cochliobolus sativus(Bipolaris sorokiniana),easy to cause the outbreak of barley leaf spot epidemic and serious economic losses under the conditions of suitable temperature,humidity and sufficient wheat root rot umbilical cord of B.sorokinianaTherefore,the most effective ways to develop breeding resistance to leaf spot and prevent leaf spot are to screen and utilize materials with persistent and stable resistance,study the mechanism of barley response to leaf spot stress and explore its resistance genes.In order to systematically research the mechanism of barley leaf spot resistance,we used two barley varieties with significant disease resistance differences(resistant variety Mengpimai 3 and sensitive variety Mengpimai 1)and strong invasive strain Z14484 as materials to systematically discuss the mechanism of barley leaf spot resistance by morphology,physiological and biochemical methods,and explore the metabolic regulation network and regulatory genes related to barley leaf spot resistance based on the transcriptomics analysis.The main findings are following:1.The resistance test and physiological index of materials in the normal growth and spray treatment were tested.It was found that the resistant variety and sensitive variety showed significant changes under the measurement period after the infection of leaf spot bacteria.The activities of lipoxygenase(LOX),malondialdehyde(MDA),soluble sugar(SS),?-1,3-glucanase and phenylalanine ammonia-lyase(PAL)in MP3 were higher than that of MP 1 at seedling stage;At jointing stage,the activities of superoxide dismutase(SOD),peroxidase(POD),catalase(CAT),polyphenol oxidase(PPO),LOX,proline(Pro),MDA,?-1,3-glucanase and PAL of MP3 were higher than that of MP 1;At booting stage,the activities of SOD,POD,PPO,LOX,Pro,SS,and ?-1,3-glucanase of MP3 increased more significantly than that of MP 1.2.Principal component analysis of 11 indicators in three measurement periods revealed LOX activity,PPO activity,POD activity,SS content,PAL activity and ?-1,3-glucanase activity had higher contribution on spot disease and were related to the tolerance of barley leaf,and which could be used as an important indicator for evaluating barley leaf spot resistance.3.The leaves of anti-disease variety MP3 inoculated for 0 h(CK,is no inoculation),3 h,6 h,12 h,24 h and 72 h were sequenced by transcriptome.The total number of expressed genes was 33,723.Among them,29191 genes were known,and 4532 new genes were predicted.A total of 54364 new transcripts were detected,of which 42972 genes belonged to the novel alternative splicing subtype of known protein-coding genes,and 4755 genes belonged to the transcription of new protein-coding genes.The remaining 6,637 genes belonged to long-chain non-coding RNAs.4.After transcription factor(TF)protein domains annotation,it found that genes were enriched in ABI3VP1,AP2-EREBP,FAR1,MYB,NAC,WRKY,bHLH and other transcription factor families;The domain types of the test genes were mostly enriched in the CNL,N,CL,RLP,TNL type domains and the RLP domain.5.The transcriptome sequencing results were used to compare and analyze the gene expression of barley at different time points with normal growth and infection of leaf spot pathogen.A total of 694 barley genes were significantly different after leaf spot disease stress.The differentially expressed genes between leaf spot-stressed barley and normal-growing barley were found in several biological pathways,including "metabolic pathways","biosynthesis of secondary metabolites" and "catalysis","active" and so on by GO,KEGG annotation.It indicated that barley was most affected by leaf spot in these pathways,which may be the main factor leading to abnormal growth of plants stressed by leaf spot..6.In this study,there were 10 genes related to enzyme metabolism by expression,GO and KEGG functional annotation,and the accuracy of transcriptome results was verified by real-time quantitative(qRT-PCR).In order to further study the key genes in transcription mechanism of barley leaf spot resistance,the relationship between enzyme metabolism and resistance gene expression that is regulated by important genes will provide a theoretical basis.