关键词: 花生丛枝植原体, 免疫膜蛋白, 细菌双杂交, 诱饵载体

Abstract: The molecular mechanisms involving insect-vector-phytoplasma-host-plant relationships were poorly understood. Our study focused on a cell-surface membrane protein of the phytoplasma named immunodominant membrane protein (Imp). The objective of experiment was to get the basic information between the mechanism of phytoplasma-host-plant relationships. The target ORF had been amplified using primers designed based on the conserved regions of imp gene from phytoplasma. The amplified gene was analyzed for phylogenetic tree, and the encoding protein was analyzed for transmembrane region, hydrophilic and hydrophobic region in this study. The imp gene was inserted into pBT vector including λcI gene. Construction of the pBT-Imp plasmid can be used for experiment followed by conformed that imp-λcI fusing protein was expressed by IPTG induction, western blot and self-validation testing. The immunodominant membrane protein gene from peanut witches’ broom (PnWB) phytoplasma was 519 bp in length, encoding a predicted protein of 172 amino acids. Homology analysis of sequence from PnWB and 17 phytoplasma showed that imp gene of PnWB phytoplasma closely related to SPWB and WBDL phytoplasma, with homology rate of nucleotide being 99.9% and 79.8%, amino acids sequence, being 100% and 70.2%. Analysis of protein structure showed that the protein from PnWB phytoplasma possesses a region with C-terminal hydrophobic transmembrane anchor and N-terminal hydrophilic characterization, possibly internal to the phytoplasma cell.

Key words: peanut witches&rsquo, broom phytoplasma;immunodominant membrane protein;Bacterio Match Ⅱ two-hybrid system;bait plasmid construction