Fenotipagem e expressão gênica para tolerância ao déficit hídrico em milho

Abstract

Understanding the mechanisms involved in water deficit tolerance is essential in maize breeding programs for the choice of safe methodologies for phenotyping. Phenotyping to water deficit tolerance in seeds, seedlings and tissues of reserve has the advantage of being performed early, with less demand for time, space and labor. In addition to phenotyping, the expression of genes in seeds and seedlings and remaining tissues of maize reserve material is important for the development of improvement strategies since the overexpression or repression of these genes influences the defense mechanisms of plants. Thus, the objective was to compare two methods to simulate water deficit conditions. Two contrasting lineages were used to water deficit tolerance. In the first method, seeds were germinated and seedlings developed in sand with 10% and 70% of water retention capacity. Characteristics related to the growth of seedlings and tissues of reserve were evaluated. In the second method, germination paper was used moistened with a PEG 6000 solution (-0.6 MPa), and with water as a control. From the third day after sowing, characters related to seed germination were evaluated. When using the sand substrate, it was possible to discriminate the lineages by the characters: shoot length, main root length, number of secondary roots and emergence at 4, 5 and 6 days and emergence speed index. In PEG 6000 solution significant interactions were observed between genotypes versus environments for the variables: germination at 4 days, emergence speed index, emergence at 4 and 7 days, root and shoot green weight, and shoot dry weight. For root dry weight, significance for environment was observed. Seeds of 203 F2:3 progenies, obtained from contrasting lineages were phenotyped for this characteristic using the PEG 6000 solution. The evaluations were carried out at five and seven days after sowing and from the data obtained and progenies were classified as tolerant, intermediate and intolerant. Of the 203 F2:3 progenies, 108 progenies with 7 days after sowing were also phenotyped by image analysis. The length of the main root and shoot and the total length were evaluated. Estimates of variance components and correlations between these two methods and between the evaluated characteristics were calculated. Correlation of 74% was observed between the data observed at 5 and 7 days and heritability values were 88.9% and 84.9%. It was concluded that phenotyping to water deficit tolerance, either through image analysis or traditionally, it can be performed five days after sowing, considering the length of the main root, and the PEG 6000 solution, to simulate water restriction, with the advantage of phenotyping by image analysis is faster and shows less labor demand. The expression of 14 genes associated with tolerance to water deficit was also evaluated by the qRT-PCR technique. For this, seedlings and remaining tissues from F2:3 progenies were used, and from tolerant and intolerant lineages, developed on germination paper moistened with PEG 6000, for 7 days after sowing. Transcription factors such as DREB 2A/2.1S, DREB2.7 and the genes related to antioxidant enzymes, CAT3, cAPX, LEA14, LEA D34 and ZmDHN1, were more expressed in the tolerant lineage and progenies. Bzip expression were associated with non-tolerance to water deficit. It is concluded that these genes can be used as markers for the early selection of maize genotypes for water deficit tolerance.