Isolation And Functional Complementation Of The Arabidopsis SDL Gene

In the previous study, short-day-dependent lesion-mimic mutant named sdl was screened by mutagenizing Columbia wild type (Col-0) with Ethyl methane sulfonate (EMS). Under long-day condition, the phenotype of the mutant has no significant difference compared with Col-0. Under short-day condition, however, small plant and random distribution of mimic lesion on leaves and inflorescences are observed. Using map-based cloning, the SDL gene was located in two BAC plasmids within 180Kb span.To isolate the SDL gene, in this experiment, sub-cloning was used. Fragments, cleaved from the two BAC using restriction enzymes, were connected into binary expression vector pCAMBIA1301, generating 35 positive clones. All these clones were transformed into Arabidopsis thanliana mutant sdl, and transgenic plants were obtained through resistance screening. After short-day treatment, there was only one kind of transgenic plant which showed the same phenotype as the wild type, with no mimic lesion observed. The other transgenic plants, however, all showed the same phenotype as sdl mutant. Sequencing result shows:the fragment from sdl mutant contains a gene with point mutation. Probably, the gene mutation leads to the formation of mimic lesion. This gene is preliminary proposed to be SDL.In order to verify the function of SDL gene, the full length sequence of SDL was amplified from Col-0 by PCR, and connected into binary expression vector pROK2. Then 16 transgenic seedlings were obtained. Under short-day condition, all the seedlings showed no mimic lesion. So it is confirmed that mutation on SDL leads to the formation of mimic lesion.SDL is an unreported gene; therefore, the isolation of SDL has scientific significance on the research of the molecular mechanism of mimic lesion formation.