Total RNA was isolated from phloem of young shoots and retro-transcripted to cDNA by RT-PCR(Reverse Transcriptase-Polymerase Chain Reaction) with specific primer pair of CVd-Ⅲ (citrus viroid-Ⅲ), the results of molec...Total RNA was isolated from phloem of young shoots and retro-transcripted to cDNA by RT-PCR(Reverse Transcriptase-Polymerase Chain Reaction) with specific primer pair of CVd-Ⅲ (citrus viroid-Ⅲ), the results of molecular detection indicated that the CVd-Ⅲ specific bands, about 300 bp, were detected in 27 of the 49 samples collected from different counties in Hunan Province and 1 sample presented by Catania University. All the amplified cDNA fragments were cloned and sequences, sequence analysis compared with the reported CVd-Ⅲ sequences in GenBank showed that: sequences of 21 samples were the same and completely identical with CVd-Ⅲb (AF184147); however, the sequences of the other 6 samples and presented sample were diversified from the reported CVd-Ⅲ sequences in GenBank.展开更多
文摘柑橘黄龙病病原菌为韧皮部限制性寄生菌Candidatus Liberibacter,迄今为止尚未成功获得离体纯培养。因此基于特异性抗体的高通量血清学检测技术迄今尚未开发。本实验室前期研究利用异源表达重组黄龙病菌外膜蛋白Omp_(CLas)免疫动物,制备并获得了特异性多克隆抗体(Anti-Omp_(CLas)-Pab)。本研究利用直接组织印迹免疫法(Direct tissue blot immunoassay,DTBIA)技术对获得的多克隆抗体的检测效果进行了初步评价,证实该抗体可以特异性识别中国不同地理来源的柑橘黄龙病菌分离物;柑橘叶柄、叶片中脉、枝条、根部、果梗和种子中均存在黄龙病菌,其中根部和种子中的病菌含量相对较低,枝条和叶片中脉中的含量较高。该多克隆抗体的获得为后续快速检测试剂盒或试纸条的研发奠定了基础。
文摘Total RNA was isolated from phloem of young shoots and retro-transcripted to cDNA by RT-PCR(Reverse Transcriptase-Polymerase Chain Reaction) with specific primer pair of CVd-Ⅲ (citrus viroid-Ⅲ), the results of molecular detection indicated that the CVd-Ⅲ specific bands, about 300 bp, were detected in 27 of the 49 samples collected from different counties in Hunan Province and 1 sample presented by Catania University. All the amplified cDNA fragments were cloned and sequences, sequence analysis compared with the reported CVd-Ⅲ sequences in GenBank showed that: sequences of 21 samples were the same and completely identical with CVd-Ⅲb (AF184147); however, the sequences of the other 6 samples and presented sample were diversified from the reported CVd-Ⅲ sequences in GenBank.
