Molecular identification of the pathogen of willow witches'-broom
To clarify the taxonomic status of the phytoplasma causing willow witches'-broom in Gansu Pro-vince,this study used nested PCR technique to clone the 16S rDNA gene using total DNA collected from differ-ent regions and species of willow witches'-broom samples as templates,and nine 1 250 bp target fragments were obtained using P1/P7 and R16F2n/R16R2;the sequences of the above genes were compared by homolo-gy,The results showed that the 16S rDNA gene of willow witches'-broom phytoplasma strain had the highest homology with Willow proliferation phytoplasma(Willow proliferation)in Inner Mongolia,both above 99.1% ;the phylogenetic tree was constructed and found that the 16S rDNA gene of willow witches'-broom clustered with members of 16SrVI-A subgroup.The phylogenetic tree was constructed,and the 16S rDNA gene of willow witches'-broom clustered with members of the 16SrVI-A subgroup;and the virtual RFLP analysis reported that the highest sequence similarity with the one obtained in this study was the reference pattern of the 16SrVI-A subgroup,with a similarity coefficient around 0.98,so the willow witches'-broom phytoplasma in Gansu Province could be classified into the 16SrVI-A subgroup.Meanwhile,the phytoplasma resources were preserved by grafting and identified as vector insects based on amplification of the 16S rDNA gene in the willow shoulder reticulata.
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