Study On Antioxidant Enzyme And Single Nucleotide Polymorphisms(Snps)Associated With Drought Stress In Brachypodium Distachyon

The severity and frequency of drought stress is unexpected to increase because of climate change, which can significantly affect on plant growth and development. Drought stress is highly variable in nature, with frequency changes in intensity and duration. And the drought is a complicated trait, and is regulated by many genes. So it is very important sutible for analysis materials and methods. Brachypodium distachyon is a temperate wild grass species and is a powerful model system for studying grain, energy, forge and turf grasses. The gene and genomic results were bases on analysis grasses. And they have large natural various. So Brachypodium distachyon was good materials for analysis drought stress. Association analysis is a good tool for the study on the relationship between genes and quantitative traits. And the accuracy was100%according to sequence directly. This research was carried out the phenotypic diversity of drought tolerance in57natural populations of Brachypodium in greenhouse for twice. Drought tolerant (T-9) and susceptible accession (B2C) were used to determine production of reactive oxygen species and antioxidative response of the leaves and roots of Brachypodium to drought stress. And5genes related to antioxidative as the candidate genes for analysis the single nucleotide polymorphisms were selected. And8peroxides genes were candidate genes according to the results of native gel analysis and MALDT-TOF-MS. Association analysis on8PODs gene SNPs and traits related to drought was present. The main results are described as followed:1. Natural variation of drought response in Brachypodium distachyonTwo experiments were conducted in a greenhouse to assess the drought tolerance of57natural populations of Brachypodium. Principle component analysis revealed that reductions in chlorophyll fluorescence (Fv/Fm) and leaf water content (LWC) under drought stress explained most of the phenotypic variation, which was used to classify the tolerant and susceptible accessions. Four groups of accessions differing in drought tolerance were identified, with3tolerant,16moderately tolerant,32susceptible and6most susceptible accessions. The tolerant group had little leaf wilting and fewer reduction in Fv/Fm and LWC, while the most susceptible groups showed severe leaf wilting and more reductions in Fv/Fm and LWC. Drought stress increased total water soluble sugar (WSS) concentration, but no differences in the increased WSS were found among different groups of accessions.2. Antioxidative metabolism of Brachypodium Distachyon under drought stressDrought stress increased the accumulation of hydrogen peroxide (H2O2) and degree of lipid pexoxidation (Malondialdehyde, MDA) in the leaves and superoxide (O2) concentration in the roots, but to a greater extent in the B2C compared with T-9. An enhanced leaf activity of glutathione peroxidase (GPX) in T-9and the decreased activities of catalase (CAT) and monodehydroascorbate reductase (MDHAR) in B2C were observed, respectively. Drought stress increased root activities of CAT and MDHAR in both accessions, to a greater degree in the tolerant T-9. Drought stress also increased root activities of GPX, ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR) and glutathione reductase (GR) in the T-9but decreased activity of superoxide dismutse (SOD) in the B2C.3. Nucleotide diversity and linkage disequilibrium in antioxidative candidate genes for drought tolerance of Brachypodium distachyonThe section of these genes was based on the significant changes in enzyme activities observed between the control and drought-stressed plants, either tolerant T-9, or susceptible B2C, or both. The average nucleotide diversity (π) for CAT, GPX, DHAR, MDHAR, and APX (encoding ascorbate peroxidase) was0.0027among accessions contrasting drought tolerance. The average single nucleotide polymorphism (SNP) frequency across these five genes was one SNP per133bβ in the total length. The linkage disequilibrium (LD) decay extended to a distance of1.6kb across all genes. The neighbor-joining tree analyses of DHAR, MDHAR, and CA T generally separated accessions differing in drought tolerance. 4. Nucleotide diversity and association analysis on POD genes and drought traitsMake sure8candidate genes through POD enzyme native gel analysis in57Brachypodium distachyon and MALDT-TOF-MS.90SNPs were from8POD genes with the sequence length9178bp. Coding part had44SNPs with the sequence length4396bp. And8genes belonged to natural test. The average indel diversity was0.652. There was557r2according to linkage disequilibrium analysis, and significant points were190and the ratio34%.The number of SNPs related with drought traits accounted for30%in90SNPs according to association analysis. There were21SNPs significantly related with drought traits in tolerance genes and accounted for70%, but the susceptible genes only have9SNPs and accounted for21%.The most related SNPs was the gene Bradilg63060.1and more related was the gene Bradi3g41340.1. There were a few related SNPs in the susceptible gene Bradilg26870.1and Bradilg65820.2and accounted for7.1%and12.5%, respectively. So the drought tolerant gene Bradilg63060.1was a good gene for molecular marker and transfer genes. The ration between transition and transversion was1.5:1based on SNP analysis results. Therefor the transition was the main base mutation.