应用多重PCR技术同步检测柑橘4种重要嫁接传播病原

 柑橘衰退病毒(Citrus tristeza virus，CTV)，柑橘碎叶病毒(Citrus tatter\|leaf virus，CTLV)，柑橘裂皮病类病毒(Citrus exocortis viroid，CEVd)和柑橘黄龙病(Huanglongbing， HLB)亚洲种病原(Candidatus liberobacter asiaticus)是重要的柑橘嫁接传播病原。本文建立了同时检测HLB病菌、CTV、CEVd 和CTLV 4种柑橘嫁接病原的一步法、双温多重PCR检测技术体系，同时在体系中设置内参基因。应用该体系快速评价了4种嫁接传播病原在田间侵染情况，结果表明28个田间样品CTV、CEVd、CTLV和HLB感染率分别为89.3 %、17.9 %、10.7 %和28.6 %，接近半数样品为混合感染。并且将该方法应用于快速评价茎尖嫁接苗病毒的脱除情况。

Simultaneous and Rapid Detection of Four Important Graft\|transmissible Pathogens of Citrus by Multiplex PCR

Citrus tristeza virus (CTV), Citrus tatter leaf virus (CTLV), Citrus exocortis viroid (CEVd), and Candidatus Liberibacter asiaticus (Ca. L. asiaticus) associated with Huanglongbing (HLB) are important graft\|transmissible pathogens of citrus. A rapid multiplex PCR protocol with an internal control was developed for the detection of these citrus pathogens using a timesaving two\|temperature cycling for PCR amplification. The assay was applied for rapid detection of those pathogens in field\|grown trees. The results indicated that 89.3%, 10.7%, 17.9% and 28.6% of the 28 field\|grown citrus samples were infected with CTV, CTLV, CEVd and Ca. L. asiaticus, respectively, and approximately half of the samples were co\|infected with more than one pathogen. In addition, the absence of multiple pathogens in shoot\|tip grafted budlings was evaluated with the multiplex PCR assay.