Two of the selected TDFs (serine/threonine-protein PLX-4720 nmr kinase and importin β) were more abundant in infected plants, whereas two TDFs (autophagy protein 5 and RNA polymerase β) showed higher expression in healthy plants. The 18 s RNA gene of Mexican lime tree was used as a reference gene for data normalization, as described previously [12]. Real-time PCR analysis showed that the expression of the selected genes agreed well with the profiles determined by cDNA-AFLP (Figure 4). Figure

4 Real-time analysis of four differentially expressed transcript derived fragments (DE-TDFs). The Y axis represents the relative expression (expression normalised to that of the housekeeping gene). Discussion In this study, we performed a comparative transcriptomic analysis of healthy Mexican lime trees and those infected by “” Ca. Phytoplasma aurantifolia”"

by using cDNA-AFLP technique. For this analysis, we used leaf samples from healthy controls and infected plants at the symptomatic stage. The symptomatic stage was chosen because the plant/pathogen interaction is well established but the plant cells are still active and can maintain pathogen survival. As far as we are aware, our study is the first gene expression analysis of the compatible interaction between “” Ca. Phytoplasma aurantifolia”" and Mexican lime trees. We observed transcriptional changes that affected the expression of several genes related to physiological functions that click here would affect most leaves in infected tissues. The cDNA-AFLP method for global transcriptional analysis is an open architecture technology that is appropriate for gene expression studies in non-model species. This is because prior sequence data are not required for the visual identification

of differentially-expressed transcripts, in contrast to other approaches. Infection with “” Ca. Phytoplasma aurantifolia”" causes widespread gene repression in Mexican lime trees Sixty-seven percent of the identified DE-TDFs were down-regulated in response to infection, Vitamin B12 whereas only 33% were up-regulated in response to infection which could reflect the exploitation of cellular resources and the suppression of defence responses by the phytoplasma [13]. Responses to external stimuli and defence Several genes that were modulated in Mexican lime trees by infection with “” Ca. Phytoplasma aurantifolia”" were related to defence, cell walls, and response to stress. The expression of autophagy protein 5 was repressed. Autophagy is a survival mechanism that protects cells against unfavourable environmental conditions, such as microbial pathogen infection, oxidative stress, nutrient starvation, and aggregation of damaged proteins [14]. It has been shown that carbohydrate starvation induces the expression of autophagy genes [15] and stimulates the formation of reactive oxidative species (ROS) in plants [14].