Characterization of tomato (Solanum lycopersicum L.) male sterile mutants putatively affected in class B MADS-box transcription factors
Author(s)
Pucci, Anna
Date Issued
June 12, 2015
Type
Doctoral Thesis
Abstract
Male sterility is defined as the inability of plants to produce or release functional pollen grains. Genic male sterility (GMS) was studied in detail in the past and used for hybrid seed production in several plant species. The hybrid production obtained using GMS is characterized by several problems that are associated with this type of male sterility. The main problem that occurs is the maintenance of the male sterile line, which involves a backcross with heterozygous maintainer plants, but the progeny will be 50% sterile and 50% fertile and selection is needed to obtain a purely male sterile seed parent. To bypass this limit, it could be helpful to use environmental-sensitive genic male sterility (EGMS) mutations, where the sterility and fertility restoration are influenced by environmental factors such as temperature and photoperiod. The EGMS was reported in many plant species and it represents a very powerful system in hybrid seed production, especially in rice. In tomato, different male sterile mutants have been described where the sterility/fertility condition is influenced by the action of temperature or photoperiod, in particular vms, pi-2, sl, TAP3 and sl-2 for the temperature and 7B-1 for the photoperiod.
This thesis aimed to the morphological and molecular characterization of EGMS genotypes in tomato. Literature data and examination of the phenotype indicated as candidates for these mutations members of the class B MADS-box transcription factors family, that specify petal and stamen organ identity. In Arabidopsis thaliana and Antirrhinum majus, there are two class B genes that, referring to the A. majus nomenclature, are known as DEFICIENS (DEF) and GLOBOSA (GLO). In tomato, as in most solanaceae, four class B MADS-box genes are found, two DEF-like members (SlDEF and SlTM6) and two GLO-like members (SlGLO1 and SlGLO2).
The main objective of the research was the phenotypic characterization of the 7B-1 mutant and the identification of the putative gene underlying the mutation. The other goals of this work were represented by studies conducted in order to characterize other tomato male sterility mutations, such as sl-2, pi-2, TAP3 and vms in environmental conditions that favor or not the expression of the sterile phenotype. The tomato 7B-1 mutant has a strong potential for production of tomato hybrid seeds, being male sterile in long days, while in short days it produces several male fertile flowers. The morphological characterization of the mutant was performed under environmental conditions that favor the sterility expression and in those that determine a partial fertility restoration. Given the phenotypic similarity between 7B-1 and sl-2, it was taken into consideration the possibility to grow this other mutant also in permissive and non-permissive conditions in order to observe the male sterile phenotype. The phenotypic analysis was aimed to verify if the mechanism that controls the sterility/fertility restoration remains unaltered changing the geographic area (Chapter 2).
All male sterile mutants described in this thesis are characterized by different alteration degrees in anther morphology and in order to understand the genetic relationship between them were carried out different allelism tests. The results indicated the allelism between 7B-1 and sl-2 and between TAP3 and pi-2 (Chapter 3).
To identify the candidate gene of 7B-1 mutation, we developed a backcross mapping population after crossing the mutant and S. pennellii. We have characterized the mapping population at molecular level using markers representing the class B MADS box genes or associated to them. The molecular screening showed that the best candidate gene to represent the 7B-1 mutation is the SlGLO2 gene localized on chr. 6. Expression analyses were performed on different flower development stages of the 7B-1 mutant and WT counterparts analyzing genes resulted differentially expressed in mutant genotype from RNA-seq analysis, plus all class B MADS-box genes and other genes known from literature as involved in the regulation network of this family. Results reported in Chapter 4 supported the idea that the candidate gene for 7B-1 mutant is a member of class B MADS-box. The
complementation experiment using the WT allele of SlGLO2 gene to transform 7B-1 plants will is ongoing and will possibly confirm that (Chapter 4).
In Chapter 5 it is reported the morphological and molecular characterization of other two male sterility mutations, pi-2 and vms, both sensitive to temperature. The phenotype showed by these two mutations is different, in the first case the flower presents a complete conversion of petals into sepals and stamens into carpels, while in the second one the abnormalities regard only the anther structure. The different phenotype suggests the involvement of different class B MADS-box genes. Molecular analysis confirms this. For pi-2 it demonstrated the involvement of SlDEF, while for vms, it was assessed the involvement of the SlGLO1 gene (Chapter 5).
Taken together, the results obtained from this thesis give new information which will be useful to elucidate the genetic and molecular basis of the male sterility mutations influenced by the action of environmental factors.
Additional information
Dottorato di ricerca in Biotecnologie vegetali
Subjects
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