Microarrays analysis suggests that phytoplasma genome expression in the plant can be modulated according to the infection stage. A real time PCR protocol was set up to study the expression profile of different 'Candidatus Phytoplasma asteris' Chrysanthemum yellows isolate (CYP) genes during infection of Arabidopsis thaliana and the two vector species Macrosteles quadripunctulatus and Euscelidius variegatus. Target genes were selected among secreted proteins, known effectors, general metabolism and specific CY genes, obtained following Illumina sequencing. Genes coding three putative secreted peptides, antigenic membrane protein (Amp), immunodominant membrane protein (Imp), four general transporters (multidrug, mechano-sensitive channel, SecY translocation component and an oligopeptide transporter), two specific transporters (arginine, Zn++), one protein involved in phospholipid metabolism and the 30S rRNA gene were selected from the CYP genome. Specific primers were designed for each selected gene, and the appropriate real time PCR protocols were optimized. Amplification efficiencies for each target were determined by amplification of five dilutions of different plasmids containing the specific amplicon. The expression level of each target gene was correlated to the actual titer of phytoplasma in each infected plant or insect sample to provide a detailed study on gene expression of an obligate parasite in the plant and insect hosts.
Risultati preliminari sull'espressione genica del fitoplasma del giallume della margherita nella pianta e nell'insetto vettore
Pacifico D;Galetto L;Abbà S;Palmano S;Rashidi M;Bosco D;
2013
Abstract
Microarrays analysis suggests that phytoplasma genome expression in the plant can be modulated according to the infection stage. A real time PCR protocol was set up to study the expression profile of different 'Candidatus Phytoplasma asteris' Chrysanthemum yellows isolate (CYP) genes during infection of Arabidopsis thaliana and the two vector species Macrosteles quadripunctulatus and Euscelidius variegatus. Target genes were selected among secreted proteins, known effectors, general metabolism and specific CY genes, obtained following Illumina sequencing. Genes coding three putative secreted peptides, antigenic membrane protein (Amp), immunodominant membrane protein (Imp), four general transporters (multidrug, mechano-sensitive channel, SecY translocation component and an oligopeptide transporter), two specific transporters (arginine, Zn++), one protein involved in phospholipid metabolism and the 30S rRNA gene were selected from the CYP genome. Specific primers were designed for each selected gene, and the appropriate real time PCR protocols were optimized. Amplification efficiencies for each target were determined by amplification of five dilutions of different plasmids containing the specific amplicon. The expression level of each target gene was correlated to the actual titer of phytoplasma in each infected plant or insect sample to provide a detailed study on gene expression of an obligate parasite in the plant and insect hosts.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
