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Screening candidate effectors from Botryosphaeria dothidea

Xiaoqiong Zhu: China Agricultural University

<div><em>Botryosphaeria dothidea</em> is an ascomycetous fungus causing apple white rot, which is a serious disease on apple in China. The genome of <em>B. dothidea</em> was sequenced and predicted to contain about 320 effector genes with less than 300 amino-acid in size and more than four cysteine residues. Fungal effectors often play essential roles in host-pathogen interactions. However, little is known about the functions of <em>B. dothidea</em> effectors. Among the selected 100 candidate effectors in this study, 30 candidate effectors (without signal peptide regions) were successfully cloned into PVX vector pGR107 and introduced into Agrobacteria tumefaciens strain GV3101 by electroporation. Potential effects of the candidate effector genes on cell death were examined using the BAX system. Candidate effector was expressed in agro-infiltrated leaves of <em>Nicotiana benthamiana</em>. At 24 h after the initial inoculation, the infiltrated leaves were challenged with A. tumefaciens cells carrying the gene of the BAX. In total, 22 candidate effectors completely inhibited programmed cell death (PCD) of <em>N. benthamiana</em> induced by the BAX gene. Expression patterns of four effectors analyzed by Reverse Transcript-quantitative PCR showed all were highly expressed from 36 to 72 h post inoculation in apple fruit. The secretional signal peptides were tested in yeast system (pSUC2 vector) and found that 10 of 12 effectors showed secretory activity. These results suggested that candidate effectors of <em>B. dothidea</em> might play important roles in its interaction with apple.</div>