Identificazione di fattori genetici che contribuiscono all'efficienza di utilizzo del manganese tramite GWAS su A.thaliana cresciuta in mancanza di manganese

The current food system must prepare itself to respond to a series of pressures such as the increase in the world population and the supply shortages caused by various types of biotic and abiotic stresses on food crops resulting from the ongoing climate change. To better exploit the available resources and obtain higher quality food, it is important to know the mechanisms of absorption, transport and accumulation of nutrients, some of which are not yet clear as in the case of Manganese. Manganese is an essential nutritional element since it plays many important roles within the cell. Both excess and lack of manganese cause negative effects on the plant leading to a reduction in the harvest. Manganese deficiency is a latent disorder, consequently it is often detected too late and corrective interventions are expensive, inefficient and imprecise on a large scale. The effects of this deficiency include the reduction of photosynthetic efficiency, the degradation of chlorophyll and the appearance of necrotic spots on the leaf surface. The study aims to identify candidate genes involved in the homeostasis of manganese, through a genomic association study (GWAS). For this study we have grown a set of 205 different ecotypes of A.thaliana in petri dishes under manganese deficiency and for each ecotype we have obtained 6 plants. Phenotypic data, in particular Fv/Fm and Mn concentration, were collected from each individual. Values of Mn concentration in the shoots were used to carry out a correlation analysis [Mn] vs (Fv/Fm) which shows a weak positive correlation (r = 0.14) between the two datasets. In addition, the Fv/Fm dataset was used phenotypic input to run a GWAS analysis. The list of candidates was then expanded by carrying out a Linkage Disequilibrium analysis with the intention to include in the results also other SNPs from the adjacent areas. We produced a table containing the names of all candidate genes, their function and the possible presence of non-synonymous mutations. Among the candidate genes identified we deepened the study of NRAMP1, which was already known to be involved in manganese uptake at root level. We performed a haplotypic analysis showing how the SNPs we have identified are associated with an increase in Fv/Fm. A real time PCR was then performed in order to measure the transcription levels of the gene, comparing ecotypes showing the highest Fv/Fm values with those showing the lowest levels. However, the results suggest that there is no significant difference in expression levels, leaving open the question about how this SNPs are responsible for the difference found in the ability of plants to photosynthesize. SNPs located in non-coding areas of the gene, such as 5'UTR, could affect protein translation as already demonstrated in several cases in plants and animals. Further studies are therefore needed to understand if and how these mutations can affect the ability to acquire and use manganese. The remaining candidate genes have been studied in depth at a bibliographic level and appear to be involved in various processes including the response to thermal stress, ABA signaling and biosynthesis of lignin. However, data from this bibliographic research do not suggest a direct relationships between these genes and the presence of Manganese. Further analyzes can then be performed using knock out mutants.