Diagnostics and Detection of Different Groups Phytoplasmas in China Using an Oligonucleotide Microarray on the Platform of ArrayTube

【作者】 王圣洁； LIN Cai-li； 严东辉； YU Shao-shuai； LI Yong； WANG Lai-fa； PIAO Chun-gen； GUO Min-wei； HUAI Wen-xia； 田国忠；

【Author】 WANG Sheng-jie;LIN Cai-li;YAN Dong-hui;YU Shao-shuai;LI Yong;WANG Lai-fa;PIAO Chun-gen;GUO Min-wei;HUAI Wen-xia;TIAN Guo-zhong;Key Laboratory of Forest Protection of State Forestry Administration,Research Institute of Forest Ecology,Environment and Protection,Chinese Academy of Forestry;

【机构】 Key Laboratory of Forest Protection of State Forestry Administration,Research Institute of Forest Ecology,Environment and Protection,Chinese Academy of Forestry；

【摘要】 The 16 S rDNA-specific oligonucleotide probes designed to detect phytoplasma have been reported.In order to find optimal specific probe and to develop the detection technique using oligonucleotide microarray on the platform of Array Tube to detect and identify phytoplasmas associated plant disease in China.PCR amplification and microarray hybridization were used to detected 15 symptomatic plants probably infected with phytoplasma and asymptomatic plants as healthy controls collected from different regions in China.Phytoplasma 16 S rDNA were detected in 13 of 15 symptomatic plants but not in all their healthy controls.Thirteen phytoplasmas were classified into 16 Sr Ⅰ,16 Sr Ⅱ,16 Sr Ⅴ and 16 Sr XIX groups.Among 17 tested probes,the universal probe designated Pp-502 could be used to detect all phytoplasmas associated with plant disease.The specific probe designated Pp Ⅰ-465 for 16 Sr Ⅰ group could be used to detected four phytoplasma strains of 16 Sr Ⅰ group associated with paulownia witches’-broom,chinaberry witches’-broom,mulberry dwarf and lettuce yellows.The probe Pp Ⅱ-629 for 16 Sr Ⅱ could be used to detect three phytoplasma strains 16 Sr Ⅱ group associated with peanut witches’-broom,sweet potato witches’-broom and cleome witches’-broom.Three phytoplasma strains of 16 Sr Ⅴ associated with jujube witches’-broom,cherry lethal yellows and Bischofia polycarpa witches broom and chestnut yellows crinkle phytoplasma of 16 Sr XIX could also be detected by specific probes,but they have obvious cross hybridization with other group probes.Compared with PCR amplification,the sensitivity of microarray to detect phytoplasma in plant was increased 1000-fold.Phytoplasmas of 16 SrI and 16 SrV group respectively were detected in periwinkle with symptoms of phyllody and witches’-broom collected from Fujian province and Robinia hispida with symptom of witches’-broom collected from Henan province.While no phytoplasma was detected in peony with symptom of yellowing collected from Beijing province and willow with symptom of witches’ broom collected from Inner Mongolia Autonomous Region.The oligonucleotide microarray on the platform of Array Tube could be used as a method to investigate phytoplasmas in China,and could provide reliable foundation for phytoplasma identification and classification.更多还原

【Abstract】 The 16 S rDNA-specific oligonucleotide probes designed to detect phytoplasma have been reported.In order to find optimal specific probe and to develop the detection technique using oligonucleotide microarray on the platform of Array Tube to detect and identify phytoplasmas associated plant disease in China.PCR amplification and microarray hybridization were used to detected 15 symptomatic plants probably infected with phytoplasma and asymptomatic plants as healthy controls collected from different regions in China.Phytoplasma 16 S rDNA were detected in 13 of 15 symptomatic plants but not in all their healthy controls.Thirteen phytoplasmas were classified into 16 Sr Ⅰ,16 Sr Ⅱ,16 Sr Ⅴ and 16 Sr XIX groups.Among 17 tested probes,the universal probe designated Pp-502 could be used to detect all phytoplasmas associated with plant disease.The specific probe designated Pp Ⅰ-465 for 16 Sr Ⅰ group could be used to detected four phytoplasma strains of 16 Sr Ⅰ group associated with paulownia witches’-broom,chinaberry witches’-broom,mulberry dwarf and lettuce yellows.The probe Pp Ⅱ-629 for 16 Sr Ⅱ could be used to detect three phytoplasma strains 16 Sr Ⅱ group associated with peanut witches’-broom,sweet potato witches’-broom and cleome witches’-broom.Three phytoplasma strains of 16 Sr Ⅴ associated with jujube witches’-broom,cherry lethal yellows and Bischofia polycarpa witches broom and chestnut yellows crinkle phytoplasma of 16 Sr XIX could also be detected by specific probes,but they have obvious cross hybridization with other group probes.Compared with PCR amplification,the sensitivity of microarray to detect phytoplasma in plant was increased 1000-fold.Phytoplasmas of 16 SrI and 16 SrV group respectively were detected in periwinkle with symptoms of phyllody and witches’-broom collected from Fujian province and Robinia hispida with symptom of witches’-broom collected from Henan province.While no phytoplasma was detected in peony with symptom of yellowing collected from Beijing province and willow with symptom of witches’ broom collected from Inner Mongolia Autonomous Region.The oligonucleotide microarray on the platform of Array Tube could be used as a method to investigate phytoplasmas in China,and could provide reliable foundation for phytoplasma identification and classification.更多还原