Spider mites (Acari: Tetranychidae) are polyphagous pests of economic importance in agriculture, among which the two-spotted spider mite Tetranychus urticae Koch has spread widely worldwide as an invasive species, posing a serious threat to fruit tree production in China, including Beijing. The hawthorn spider mite, Amphitetranychus viennensis Zacher, is also a worldwide pest of fruit trees and woody ornamental plants. The cassava mite, Tetranychus truncatus Ehara, is mainly found in Asian countries, including China, Korea and Japan, and mainly affects fruit trees and agricultural crops. These three species of spider mites are widespread and serious fruit tree pests in Beijing. Rapid and accurate identification of spider mites is essential for effective pest and plant quarantine in Beijing orchard fields. The identification of spider mite species is difficult due to their limited morphological characteristics. Although the identification of insect and mite species based on PCR and real-time polymerase chain reaction TaqMan is becoming increasingly common, DNA extraction is difficult, expensive and time-consuming due to the minute size of spider mites. Therefore, the objective of this study was to establish a direct multiplex PCR method for the simultaneous identification of three common species of spider mites in orchards, A. viennensis, T. truncatus and T. urticae, to provide technical support for the differentiation of spider mite species and phytosanitary measures in orchards in Beijing. Based on the mitochondrial cytochrome c oxidase subunit I (COI) of the two-spotted spider mite and the cassava mite and the 18S gene sequence of the hawthorn spider mite as the amplification target, three pairs of specific primers were designed, and the primer concentrations were optimized to establish a direct multiplex PCR system for the rapid and accurate discrimination of the three spider mites without the need for DNA extraction and purification. The method showed a high sensitivity of 0.047 ng for T. truncatus and T. urticae DNA and 0.0002 ng for A. viennensis. This method eliminates the DNA extraction and sequencing procedures of spider mite samples, offers a possibility for rapid monitoring of multiple spider mites in an integrated microarray laboratory system, reducing the time and cost of leaf mite identification and quarantine monitoring in the field.

BACKGROUND:Jujube is an economically important fruit tree and native to China. Viral disease is a new threat to jujube production, and several new viruses have been identified infecting jujube plants. During our field survey, jujube mosaic disease was widely distributed in Beijing, but the associated causal agents are still unknown.METHODS:Small RNA deep sequencing was conducted to identify the candidate viruses associated with jujube mosaic. Further complete genome sequences of the viruses were cloned, and the genomic characterization of each virus was analyzed. The field distribution of these viruses was further explored with PCR/RT-PCR detection of field samples.RESULTS:Mixed infection of four viruses was identified in a plant sample with the symptom of mosaic and leaf twisting, including the previously reported jujube yellow mottle-associated virus (JYMaV), persimmon ampelovirus (PAmpV), a new badnavirus tentatively named jujube-associated badnavirus (JaBV), and a new secovirus tentatively named jujube-associated secovirus (JaSV). PAmpV-jujube was 14,093 nt in length with seven putative open reading frames (ORFs) and shared highest (79.4%) nucleotide (nt) sequence identity with PAmpV PBs3. Recombination analysis showed that PAmpV-jujube was a recombinant originating from plum bark necrosis stem pitting-associated virus isolates nanjing (KC590347) and bark (EF546442). JaBV was 6449 bp in length with conserved genomic organization typical of badnaviruses. The conserved RT and RNAse H region shared highest 67.6% nt sequence identity with jujube mosaic-associated virus, which was below the 80% nt sequence identity value used as the species demarcation threshold in Badnavirus. The genome of JaSV composed of two RNA molecules of 5878 and 3337 nts in length, excluding the polyA tails. Each genome segment contained one large ORF that shared homology and phylogenetic identity with members of the family Secoviridae. Field survey showed JYMaV and JaBV were widely distributed in jujube trees in Beijing.CONCLUSION:Two new viruses were identified from jujube plants, and mixed infections of JYMaV and JaBV were common in jujube in Beijing.

[目的]探讨片突菱纹叶蝉Hishimonus lamellatus Cai et Kuo不同种群中枣疯植原体与沃尔巴克氏体Wolbachia的感染情况和Wolbachia在不同器官组织分布,明确枣园菱纹叶蝉中Wolbachia的感染类型和分类地位,为研究Wolbachia感染对枣疯植原体潜在介体叶蝉生物学及生态学影响奠定基础.[方法]通过枣疯植原体和Wolbachia的基因特异性引物对片突菱纹叶蝉田间自然种群和实验室种群进行分子检测和鉴定.[结果]田间采集的片突菱纹叶蝉成虫植原体感染率在55％-61％之间,而Wolbachia感染率为3％-4％.田间采集的片突菱纹叶蝉自然种群经室内饲养,在1-4龄若虫中检测到Wolbachia,2-5龄若虫中检测到了植原体.片突菱纹叶蝉实验室饲养无植原体种群在其卵巢、卵和若虫中发现感染Wolbachia,在其唾液腺和消化道也检测到了Wolbachia,感染率在58％-100％之间.基于Wolbachia的wsp基因构建系统发育树,发现片突菱纹叶蝉体内的2个Wolbachia株系同属于B大组,但不同于B大组其他株系,属于新株系wLam1和wLam2.[结论]片突菱纹叶蝉成虫采自田间种群可以感染枣疯植原体和Wolbachia,无植原体叶蝉实验室饲养种群成虫感染Wolbachia显著高于田间种群,片突菱纹叶蝉体内2个Wolbachia株系属于B大组.这一研究结果为Wolbachia作为介体叶蝉生物防治剂进一步利用提供了基础信息.

为了探索松毛虫赤眼蜂防治高粱田棉铃虫、玉米螟和桃蛀螟技术,保护和利用天敌昆虫,减少化学农药使用,于2019年在高粱田人工释放松毛虫赤眼蜂,每次1.5万头/亩,卵卡挂在高粱植株上部叶片背面.同时以高氯甲维盐防治作为对照,利用性信息素诱捕器诱杀上述3种鳞翅目害虫雄蛾,定期调查记录3种害虫和主要捕食性天敌昆虫的数量和高粱受害情况.示范结果表明,施药区和放蜂区3种害虫种群数量均下降,天敌种群数量动态变化差异较大,高粱植株受害率差异显著.高粱田放蜂后对3种害虫种群数量均在1个月后显现明显抑制作用.施药区与放蜂区害虫天敌种群数量存在显著差异,施药区捕食性天敌昆虫如瓢虫、草蛉、食虫虻和蜘蛛种群数量均显著低于放蜂区.放蜂区高粱百株叶片被害株率低于11.5％;折叶率稳定在3.5％,蛀叶脉率达到11％,蛀茎率最高为4.5％,穗被害率最高达2.5％,均显著低于施药区.说明用松毛虫赤眼蜂防治高粱田3种主要鳞翅目害虫,可显著降低高粱受害程度,达到减少化学农药使用的效果.

[目的]探究6个枣树品种枣疯病抗性差异以及枣树体内植原体迁移特性差异,为抗枣疯病枣树品种的选择利用以及枣疯病的防治提供基础数据.[方法]通过采集携带植原体的鸡心脆枣接穗,嫁接到健康枣树砧木上,采用田间调查与普通PCR技术相结合方法,比较不同品种抗枣疯病情况与植原体在不同枣树品种中迁移情况.[结果]不同枣树品种发病时间不同,发病率差异显著,蜂蜜罐枣、尜尜枣和猴头枣为抗病类型,冬枣、红螺脆枣和鸡心脆枣为感病类型.植原体在不同枣树品种中迁移速度不同,植原体在鸡心脆枣和冬枣内迁移速度最快,其后依次为蜂蜜罐枣、红螺脆枣、尜尜枣,在猴头枣内迁移速度最慢.[结论]6个枣树品种中蜂蜜罐枣、尜尜枣和猴头枣表现一定的抗病能力,初步确认了植原体在不同枣树品种体内迁移特征的差异,可为抗枣疯病品种的选择利用和枣疯病防治提供科学依据.

[目的]为了解枣疯植原体免疫优势膜蛋白的特征,并进行初步表达.[方法]对北京农学院农业农村部华北都市农业重点实验室测得的枣疯植原体JWB-Dongzao免疫优势膜蛋白基因imp-DZ序列进行一致性、进化树和蛋白信号肽与跨膜区分析,分别设计引物扩增完整imp-DZ序列和去除跨膜区编码序列,克隆到蛋白表达载体pBM30上进行原核表达.[结果]imp-DZ基因序列大小为459 bp,编码152个氨基酸,除与枣疯植原体Jwb-nky中一个编码假定蛋白的基因一致性达到96％之外,与其他植原体imp基因核苷酸和氨基酸序列一致性均低60％.枣疯植原体JWB-Dongzao的imp-DZ与16SrV组其他成员imp基因聚在一个分支.携带Imp-DZ跨膜区的重组菌无法表达该蛋白,去掉跨膜区后蛋白得到大量表达.[结论]该研究成功表达了枣疯植原体的免疫优势膜蛋白Imp-DZ,证实了跨膜区影响其在大肠杆菌中的表达,为下一步制备该蛋白抗血清和探索植原体与寄主植物互作机制提供依据.

为进一步研究虫酰肼在苹果树冠层的递释规律,分别采用QuEChERS法和Florisil固相萃取法建立虫酰肼在苹果叶片上的液相分析检测方法.结果 显示,QuEChERS法虫酰肼的均添加回收率为80.34％-99.20％,变异系数为5.3％-17.9％,最小检出量均为0.1 ng,最低检出质量浓度均为0.01 mg· L-1;Florisil固相萃取法虫酰肼的均添加回收率为78.45％-88.24％,变异系数为7.8％-10.9％,最小检出量均为0.1ng,最低检出质量浓度均为0.02 mg· L-1.两种方法均满足农药残留分析的要求.

During the summer of 2018, one Syringa reticulata plant showing witches' broom and small leaves was observed in Beijing, China. Molecular diagnostic tools and electron microscopic cell observation were used to detect the possible pathogen of this disease. As a result, the phytoplasma in the symptomatic S. reticulata tree was confirmed by amplifying the 16S rRNA gene using the phytoplasma-specific universal primer pair R16mF2/R16mR1 and observation with transmission electron microscopy. The rp and tuf genes of the phytoplasma were also cloned and sequenced as the 16S rRNA gene. Sequence and phylogenetic analyses of the 16S rRNA, rp and tuf genes indicated that the phytoplasma associated with S. reticulata witches' broom (SrWB) disease belonged to the 16SrV-B subgroup, and it was closely related to the 16SrV-B subgroup phytoplasma strain jujube witches' broom, which causes serious disease of jujube fruit trees in China. This study shows the S. reticulata tree as a new host of a phytoplasma belonging to the 16SrV-B subgroup in China.

In recent years, we found that Hishimonus lamellatus Cai et Kuoh is a potential vector of jujube witches'-broom phytoplasma. However, little is known about the anatomy and histology of this leafhopper. Here, we examined histology and ultrastructure of the digestive system of H. lamellatus, both by dissecting and by semi-and ultrathin sectioning techniques. We found that the H. lamellatus digestive tract consists of an esophagus, a filter chamber, a conical midgut and midgut loop, Malpighian tubules, an ileum, and a rectum. Furthermore, both the basal region of the filter chamber epithelium and the apical surface of the midgut epithelium have developed microvilli. We also identify the perimicrovillar membrane, which ensheaths the microvilli of midgut loop enterocyte, and the flamelike luminal membrane, which covers the microvilli of the conical midgut epithelium. In addition, H. lamellatus has the principal and accessory salivary glands. Our observations also showed that the endoplasmic reticulum, mitochondria, and secretory granules were all highly abundant in the secretory cells of the principal salivary glands, while the accessory glands consist of only one ovate or elbow-like acinus. We also briefly contrast the structure of the gut of H. lamellatus with those of other leafhopper species. These results intend to offer help for the future study on the histological and subcellular levels of phytopathogen-leafhopper relationships, including transmission barriers and the binding sites of pathogens and other microorganisms within their leafhopper vectors.

[目的]筛查北京地区枣园感染枣疯病植原体的叶蝉种类,确定枣疯病植原体潜在介体昆虫,为预防和控制枣疯病流行提供科学依据.[方法]通过扫网法和黄板诱集法,对昌平区流村镇流村、海淀区西北旺镇唐家岭村、朝阳区奥林匹克森林公园、通州区永于路北京观光南瓜园4个地点枣树上叶蝉种类及枣疯病植原体感染的叶蝉进行调查和分子检测分析.[结果]北京地区枣园枣树上有13种叶蝉发生,其中包括片突菱纹叶蝉(Hishimonus lamellatus Cai et Kuoh)、凹缘菱纹叶蝉(H.sellatus (Uhler》)、小绿叶蝉(Empoasca spp.)、斑叶蝉(Erythroneura sp.)、横带叶蝉(Sca phoideus festivus Matsumura)、大青叶蝉(Cicadella viridis (Linnaeus))、新县长突叶蝉(Batracomorphus xinxianensis Cai et Shen)、红闪小叶蝉(Zygina sp.)、白边大叶蝉(Kolla paulula (Walker》)、桃一点叶蝉(Singapora shinshana(Matsumura))、一点木叶蝉(Phlogotettix cyclops (Mulsant et Rey》)、条沙叶蝉(Psammotettix striatus (Linnaeus》)和窗耳叶蝉(Ledra auditura Walker).通过对采集的叶蝉标本采用植原体通用引物进行PCR检测,片突菱纹叶蝉(H.lamellatus)、凹缘菱纹叶蝉(H.sellatus)、大青叶蝉(C.viridis)、白边大叶蝉(K.paulula),均发现感染枣疯病植原体,感染率分别为1.4％、3.0％、7.4％、8.3％,其他叶蝉经检测未发现感染植原体.[结论]凹缘菱纹叶蝉(H.sellatus)、片突菱纹叶蝉(H.lamellatus)、大青叶蝉(C.viridis)、白边大叶蝉(K.paulula)可能为潜在的枣疯病植原体介体昆虫.

Residues and dietary risk assessment of tetraconazole and bifenazate were investigated in strawberry under greenhouse conditions using high performance liquid chromatography (HPLC-DAD) after QuEChERs extraction in Beijing of China. The effects of different processing factors on the two pesticides were studied. The recoveries of tetraconazole and bifenazate were 87.0% and 89.1%, respectively. The dissipation curves of tetraconazole and bifenazate were in accordance with the first-order kinetic equation and the half-life were 5.92 d and 5.58 d, respectively. When the pre-harvest interval (PHI) was 3 d, the risk quotient (RQs) of both pesticides was less than 100%. Although soaking was a poor way to remove the two pesticides and heating at high temperatures increases the concentration of both pesticides, the residues of two pesticides can be effectively removed by washing after soaking. The results of dietary intake assessment indicated that potential dietary risk caused by tetraconazole and bifenazate in strawberry were acceptable for Chinese consumers.

去甲斑蝥素(NCTD)是斑蝥素(CTD)的衍生物,可人工合成.去甲斑蝥素对害虫有毒杀作用.为明确它是否有拒食作用及其作用方式,本研究以小菜蛾幼虫为供试昆虫,采用叶碟浸液法对试虫拒食作用及取食量、取食选择、取食次数和持续时间等取食行为进行了生物测定.拒食作用结果显示,2-48 h内NCTD处理组的幼虫取食量明显低于对照组且两者差距随时间逐渐增加,拒食率与剂量呈显著正相关,增加剂量会增加拒食率.行为研究结果显示,小菜蛾处理组试虫的单次平均取食时长明显减少,仅为对照组的70.6％,而持续取食次数和试探次数均明显增多,分别是对照组的1.4倍和1.9倍.结果表明,NCTD对小菜蛾幼虫的拒食作用主要表现在降低试虫的取食量,减少取食时间,增加试探次数.

To investigate the insecticidal mechanism of cantharidin, a promising biological pesticide substance from blister beetle, on Sf9 cells, a cultured cell line derived from fall armyworm, Spodoptera frugiperda, we preliminary studied the attribution of Bax channel and mitochondrial permeability transition pore on cantharidin-induced mitochondrial apoptosis signal pathway. Changes in cell morphology, activity of mitochondrial dehydrogenases, release of cytochrome C and mitochondrial transmembrane potential were detected when the two channels were blocked by specific inhibitors, Bax channel blocker and cyclosporin A. Results showed that cantharidin-induced apoptotic features, including changes in the cell morphology, release of cytochrome C and decrease in mitochondrial transmembrane potential could be significantly inhibited by Bax channel blocker, while cyclosporin A accelerated the downward trend of mitochondrial dehydrogenases activity and caused a decrease of Ca2+ in mitochondria. In summary, Bax might be necessary but not exclusively for the apoptosis induced by cantharidin and the attribution of these channels seems to be more complexity.

The insect family Cicadellidae includes economically important vectors of plant pathogens. Hishimonus sellatus (Uhler) transmits jujube witches'-broom (JWB). Currently, H. sellatus and Hishimonus lamellatus Cai et Kuoh are observed to co-occur at the same locality on jujube. H. lamellatus is now suspected to be a JWB vector. As such, correct identification of Hishimonus species present in vineyards is essential for epidemiological surveys. However, traditional identification of Hishimonus by morphology is limited to the adult male. We provide a comprehensive description of morphological and molecular tools for discriminating between H. sellatus and H. lamellatus, for use in identification and monitoring of the two Hishimonus species and studies of their plant hosts. A rapid and inexpensive method is introduced to identify H. sellatus and H. lamellatus occurring in jujube orchards. This method is based on amplification of mitochondrial cytochrome oxidase I (COI) gene, using PCR with multiplexed, species-specific primers. The reliability of this new method has been tested on different populations from different sites in Beijing region of China.

[目的]目前发现,北京枣园中的凹缘菱纹叶蝉Hishimonus sellatus (Uhler)和片突菱纹叶蝉Hishimonus lamellatus Cai混同发生.已知凹缘菱纹叶蝉可以传播枣疯病,而片突菱纹叶蝉是否携带枣疯病植原体尚待证明.正确鉴别区分枣园中菱纹叶蝉的种类并测定其体内感染枣疯病植原体情况有助于阐明田间枣疯病的流行规律,从而提出有效的预防枣疯病及其媒介昆虫措施显得十分重要.传统形态学鉴定两种菱纹叶蝉种类的方法局限于雄性成虫外生殖器,本研究目的在于建立一种快速的分子生物学方法,在区分枣园中两种枣菱纹叶蝉的同时,可检测虫体内的枣疯病植原体.[方法]以凹缘菱纹叶蝉和片突菱纹叶蝉的COI基因以及枣疯病植原体的16S rDNA为扩增目标,分别设计引物,建立一种包含3对引物的多重PCR体系.测试该多重PCR体系对叶蝉总DNA的灵敏度、准确性,以及当两种叶蝉DNA同时存在时的辨别能力和对枣疯病植原体16S rDNA的灵敏度.[结果]该多重PCR可以准确区分凹缘菱纹叶蝉和片突菱纹叶蝉,并对虫体内枣疯病植原体实现检测,其对昆虫总DNA的灵敏度达到0.012 ng,对枣疯病植原体16S rDNA模板的灵敏度达到900拷贝.[结论]该方法极大方便了对枣菱纹叶蝉的田间种群发生动态及虫体中枣疯病植原体感染的监测.

The brabybaenid land snail, Cathaica fasciola (Draparnaud, 1801) is widely distributed in China and causes significant damage to economic crops but there have been few ecological studies on this pest species. In the present study, the effects of availability of moisture and its interaction with live weight of dormant C. fasciola were examined in the laboratory for effects on the length of time that snails required to resume activity. Snail live weight increased with availability of water during transition of dormancy to activity. The amount of water uptake required to break dormancy, proportional to snail live weight, was less for larger snails than for smaller snails. The duration of dormancy breaking was much longer at a water temperature of 5 °C than at 10–30 °C. These results are discussed in relation to prospects for control of C. fasciola in crops by manipulating dormancy.

分别以感染百合无症病毒、黄瓜花叶病毒和百合斑驳病毒的百合及感染烟草花叶病毒和马铃薯病毒Y的烟草为试材,根据5种植物病毒外壳蛋白基因保守序列设计引物和寡核苷酸探针,并制备基因芯片.用Trizol试剂盒提取感染病毒的植物总RNA,荧光RT-PCR产物与芯片杂交,研究PCR产物是否进行变性处理、杂交时间、杂交温度、杂交液组分SSC和SDS浓度及PCR体系中非荧光引物和荧光引物比例对芯片杂交的影响.结果表明:杂交适宜条件为6×SSC、0.2％ SDS的杂交液、42℃杂交60 min,PCR体系中非荧光与荧光引物比例为1:10,PCR产物要进行变性处理.经过整体条件优化后的基因芯片在杂交检测上具有较高的特异性,适于检测百合病毒病.

[目的]为了更好地了解性信息素结合蛋白(pheromone binding proteins,PBPs)在桃蛀螟Conogethes punctiferalis (Guenée)嗅觉识别过程中的作用,明确其与配体化合物的结合特性.[方法]本研究利用RT-PCR结合RACE方法克隆了桃蛀螟一个性信息素结合蛋白基因;采用Real-time PCR方法分析了该蛋白在桃蛀螟不同发育阶段及雌雄蛾间的表达差异;利用荧光竞争结合实验对Cpun-PBP1蛋白与16种配基化合物的结合特性进行了分析.[结果]克隆了一个桃蛀螟性信息素结合蛋白基因,命名为Cpun-PBP1(GenBank登录号:KP027486).Cpun-PBP1开放阅读框全长510 bp,编码169个氨基酸,预测分子量为19.12 kDa,等电点为5.09,N-末端包括由起始位置开始的30个氨基酸组成的信号肽.蛋白特征分析显示,该氨基酸序列具有昆虫气味结合蛋白的典型特征,即含有6个保守的半胱氨酸残基.Cpun-PBP1在桃蛀螟成虫阶段表达量最高,且几乎全部在触角中表达,卵期微量表达,幼虫期和蛹期均不表达.通过构建Cpun-PBP1原核表达载体,诱导并获得Cpun-PBP1重组蛋白.荧光竞争结合实验对2种性信息素组分和14种寄主植物挥发物的结合力发现,Cpun-PBP1不但能有效地与桃蛀螟性信息素组分(顺-10-十六碳烯醛和十六醛)结合,结合常数分别为7.32和9.39 μmol/L;还能与8种寄主植物挥发物有效结合;其中,与莰烯的结合能力最强,结合常数为3.76 μmol/L.[结论]根据这些结果,我们推测Cpun-PBP1在桃蛀螟感受性信息素和寄主植物挥发物的过程中发挥着双重作用.

条华蜗牛近年来在北京地区多种植物上普遍发生。为了给有效防治该蜗牛提供参考依据，研究其对多种植物的取食选择性。结果表明，条华蜗牛对25种植物取食差异明显，在测试的25种植物中，仅取食其中的12种植物，取食量随蜗牛体重增加有增加趋势，并与蜗牛体重正相关。在取食的12种植物中，取食率低于27．00％的植物有7种。除体重组Ⅴ的蜗牛对旱柳取食量显著高于菠菜、龙葵和虎耳草的外，蜗牛取食量在这7种植物及各体重组之间都无显著差异（P＜0．05）。取食率高于60％的植物有5种，其取食率和取食量都极显著高于其他7种植物（P＜0．01）。体重组Ⅴ与Ⅳ的蜗牛对国槐、桃、一串红的取食量差异极显著，且都极显著高于体重组Ⅰ和Ⅱ。体重组Ⅲ的蜗牛对国槐、一串红和生菜的取食量极显著高于体重组Ⅰ和Ⅱ。蜗牛对生菜、国槐、牵牛的取食量在体重组Ⅰ和Ⅱ之间差异极显著；对牵牛的取食量在各体重组之间差异显著。各体重组的蜗牛对牵牛和生菜的取食量差异不显著，但都极显著高于其他植物。体重组Ⅲ～Ⅴ的蜗牛对桃的取食量极显著高于国槐和一串红。

分离纯化枣疯病植原体基因组DNA,有助于进一步开展对此病原全基因组测序的研究,CTAB法提取枣疯病叶片的总DNA后,采用氯化铯双苯酰亚胺密度梯度离心法从枣树总DNA中富集纯化枣疯病植原体DNA,并通过Real-time PCR方法对分离纯化效果进行定量检测.在此基础上,探索了不同CsCl初始密度对DNA分离效果的影响.在20℃,初始密度为1.650 0 g/cm3,经206 000×g下离心23 h后,感染枣疯病样品(IS)的离心管中出现2条DNA亮带,正常枣树样品(NS)离心管中只有1条.Real-time PCR检测结果表明NS管中的条带为枣树基因组DNA;IS管中与NS管中相同位置的条带为枣树基因组DNA,另1条带为枣疯病植原体基因组DNA.在保留其他试验条件下,不同CsCl初始密度,会影响DNA条带的位置,也会影响枣疯病植原体DNA与枣树DNA的分离效果,在1.562 2 g/cm3的初始浓度下分离效果最好.采用超速离心法可以有效地从感染枣疯病的枣树中分离得到纯的枣疯病植原体DNA,同时利用Real-time PCR法可以实现分离效果的评价,利用此方法分离得到的DNA可用于枣疯病植原体的全基因组测序.