Regeneration and selection of quince ba29 (cydonia oblonga mill.) somaclones tolerant to lime-induced chlorosis

Quince is the most used rootstock in Italian pear orchards, but due to the its high susceptibility to calcareous soils it induces widespread chlorosis problems on the grafted pear cultivars. Until recently, the common breeding techniques have not allowed the simple production of tolerant quince selections. Thus, somaclonal variation, and the large genetic variability brought about during the in vitro processes, might be a suitable tool to obtain genotypes with better agronomic characteristics. The aim of the present research was to induce in vitro regeneration of quince shoots under Fe-limiting conditions from leaves (of quince clone 'BA29'), and subsequently to evaluate their tolerance to lime-induced chlorosis. Leaves of explants were placed onto MS medium supplemented with TDZ and NAA, and the selective pressure conditions were obtained by reducing the normal MS iron content and/or by adding different concentrations of potassium bicarbonate (KHCO3). Selective conditions decreased both the percentage of shoot-forming leaves and the mean number of shoots per organogenetic leaf, as the iron content was reduced and/or the KHCO3 content was increased. A discrete number of putative somaclones, showing a range of severe chlorosis symptoms compared to a wild type clone, were chosen by visual assessments, and further tests were conducted under reduced amounts of Fe and the presence of 1.0 mM of KHCO3 conditions. DB-34 somaclone seemed to be the most promising one, because its chlorophyll content did not reduce as happened in the other somaclones and with the wild type clone. Stress conditions affected only the Fe2+ content of the DB-43 somaclone, even if under normal conditions the Fe2+ content was much higher than with all the other accessions. On the contrary, the medium acidification capacity of all somaclones was influenced similarly by treatments. A preliminary in vivo test confirmed these results. Moreover, preliminary RAPDs analyses disclosed polymorphism within the somaclones and among them and the wt clone. The sequence of chloroplastic protein D1, codified by the psbA gene, was determined in quince 'BA29' and pear 'Conference' to predict its hypothetical functions. Sequences showed differences in length and in amino acidic composition. In quince, the main modification is located in the '191' position where tyrosine substitutes histidine, causing a loss of one of the iron-binding sites.