Characeration And Analysis Of Wax Mutant Induced By EMS In Barley

Drought is one of the most important factors limiting the production in barley(Hordeum vulgare L.). Responsing to drought stress of crop is a complexphysiological and biochemical process, involving many genes‘expression andregulation. Cuticular wax plays an important role in the process of crop droughtresistance, but genes and their functions related to the wax biosynthesis and transferare still unknown, limiting drought resistance breeding with a goal of wax. Amutagenesis library was constructed by inducing with EMS and two non-waxymutants were selected. Phenotype of one mutant was characterized and thedifferentially expressed genes between mutant and wild type were analyzed by digitalgene expression profiling, providing useful information for the research of waxsynthesis and transport. The main results are as follows:1. After induced by EMS, many mutations were found in the M1population fromGanpi6and08-DM-04, such as yellowing, dwarf, spike branching, infertility,semi-sterility, and so on. In the M2population, some mutants with albino,yellowing, dwarf, less tillers, dense panicle, infertility, semi-male sterility andnon-wax were found. Many mutats appeared in the M1plants did not pass on tothe next generation of M2. Leaf mutant (yellowing, bleaching, etc.) affected thenormal growth of plants, and their M3generations were not obtained. The mutantlack of wax on the epidermis of leaf sheath and lemma may pass on to the M3generation.2. Compared with wild type, there was no obvious wax crystal layer on the leafsheath and lemma of mutant MB-1by scanning electron microscope, but noobvious difference on the leaf surface of them. As delay of the drought stress time,wild type leaf sheath wax content showed a trend of decline after rising first, andthe leaf mutant waxy content almost inconvenience, maintaining a low level. Waxplays an important role for water loss of plants. The water loss rate of wild typeswith wax on leaf sheath was lower than that of mutants without wax. Thereciprocal cross result showed that the mutant gene, resulting in non-wax on leafsheaths, was recessive and in nuclear, and the gene, resulting in non-wax on lemmas, was dominant and in nuclear. These two genes will be mapped using theseparation population constructed.3. Based on gene expression analysis between mutation and wild type under normalirrigation and four days‘drought stress condition by digital gene expressionprofiling (DGE), normal irrigation DGEs M-A and WT-A and drought stressDGEs M-B and WT-B were obtained.19223,19208,19420and19496genes weredetected in samples of M-A, M-B, WT-A and WT-B, respectively. Under normalirrigation condition,429genes were differentially expressed between the mutatand wild type. Among these gens,200genes were up-regulated, and229weredown-regulated. After4days‘drought stress,484differentially expressed geneswere detected between mutat and wild type,171genes up-regulated and313genesdown-regulated.4. According to the result of clustering of gene expression patterns, genes in thesame physiological and biochemical reaction would be clustered together. Thefunctions of all differentially expressed genes were annotated by GO functionanalysis. By the significance analysis of KEGG pathway, differentially expressedgenes functioned in the main metabolism and signal transduction pathways weredetermined. In cutin, suberine and wax biosynthesis pathway, five genesdifferentially expressed in wax synthesis pathway, among which two were theCER1genes and three were FAR genes. In fatty acid elongation pathway, threeKCS (beta ketoacyl-CoA synthase) genes differentially expressed were detected.