文献类型：期刊文献

中文题名：实时荧光定量PCR(SYBR Green I)检测不同抗枣疯病枣树品种嫁接接穗中的植原体浓度

英文题名：A real-tim e( SYBR Green I) PCR assay for detection and quantification of jujube w itches'-broom phytoplasm a in the grafted jujube cultivar scions w ith different resistance

作者：任争光[1] 王合[2] 林彩丽[1] 刘曦[2] 宋传生[1] 冯术快[3] 于少帅[1] 卢绪利[3] 田国忠[1]

第一作者：任争光

机构：[1]中国林业科学研究院森林生态环境与保护研究所,国家林业局森林保护学重点实验室,北京100091;[2]北京市林业保护站,北京100029;[3]北京市昌平区园林绿化局,北京102200

年份：2015

卷号：45

期号：5

起止页码：520-529

中文期刊名：植物病理学报

外文期刊名：Acta Phytopathologica Sinica

收录：CSTPCD;;北大核心:【北大核心2014】；CSCD:【CSCD2015_2016】；

基金：国家“863”计划(2012AA101501);北京市林业科学技术推广项目[京园绿推(2011)03号]

语种：中文

中文关键词：枣疯病;植原体;抗病品种;实时荧光定量PCR;嫁接传病

外文关键词：Jujube witches ＇ -broom; phytoplasma; resistant cultivar; real-time quantitative PCR; graft transmission of disease

分类号：S432.4

摘要：在枣疯植原体16S rDNA片段内设计并合成引物对DZ16SF-2/DZ16SR-2（扩增190 bp片段）,用重组质粒pMD18-DZ16S为模板建立标准曲线,并建立枣疯植原体实时荧光定量PCR（SYBR Green I）检测体系。溶解曲线分析显示：该引物特异性好,该体系对质粒模板的检测灵敏度达102 copies/μL。利用该体系对不同月份、不同发病级别的嫁接接种枣树接穗样品进行植原体含量测定。通过对高抗品种‘星光’,中抗品种‘黑腰子枣’、‘嘎嘎枣’和‘葫芦枣’,感病品种‘月光枣’、‘马牙枣’、‘莲蓬子’、‘北流西冬枣’和高感品种‘大红袍枣’嫁接到发病‘冬枣’砧木上所表现出的不同病级、病情指数和病原浓度比较结果发现,表现无症状的样品中植原体的含量在每克鲜重10^5～10^6个,发病Ⅰ～Ⅱ级枝条中植原体浓度在10^6～10^7个/g,Ⅲ～Ⅳ级发病枝条含量在10^7～10^8个/g鲜重,而V级发病枝条达到每克鲜重109个,表明接种发病的接穗症状严重度与其体内植原体浓度积累密切相关。高抗枣疯病‘星光’品种嫁接接穗体内病原浓度明显低于感病品种,而且有较多‘星光’接穗嫁接的发病砧木症状有所减轻,植原体浓度也有所降低。本研究可为探索枣树品种的抗病机制、抗病性鉴定及枣疯病植原体的防治提供参考。
Jujube witches’-broom （JWB） disease is one of the destructive diseases in Jujube fruit tree. In this study, primer pair DZ16SF-2/DZ16SR-2 was designed and synthesized, which could amplify 190 bp fragments of 16S rDNA sequence of JWB phytoplasma by PCR. The real-time （SYBR Green I） quantitative PCR detection system was constructed by using the recombinant plasmid of pMD18-DZ16S as the template for establishing standard curve. All the results showed that the primer pair DZ16SF-2/DZ16SR-2 had good specificity and sensitivity for the detection of JWB phytoplasma. The phytoplasma concentrations as well as disease index were quantitively detected and compared in the graft inoculated different cultivar scions with different grades of disease including the JWB-highly-resistant cultivar ‘Xingguang’; moderate resistant cv. ‘Heiyaozizao’, ‘Gagazao’ and ‘Huluzao’; susceptible cv. ‘Yueguangzao’, ‘Mayazao’, ‘Lianpengzizao’ and ‘Beiliuxidongzao’ and the highly susceptible cv. ‘Dahongpaozao’. In general, the phytoplasma concentrations in the asymptomatic scions were about 10^5-10^6 cells per gram of plant tissues, 10^6-10^7 and 10^7-10^8 cells per gram of tissues in the grade Ⅰ～Ⅱ and Ⅲ～Ⅳ infected scions respectively, and up to 109 per gram of tissues in the most severe grade V diseased scions, which suggested that the degree of infection and phytoplasma accumulation in different jujube scions was closely correlated with the level of visual witches’-broom symptoms. The results of the detailed investigation and detection of the JWB-highly-resistant cultivar ‘Xingguang’ indicated that the phytoplasma concentrations in the infected scions were obviously lower than other susceptible cultivars, furthermore, the grafting of the ‘Xingguang’ scions caused a reduction in the symptom severity and phytoplasma concentration in the rootstocks. This work will give the hands of exploring the jujube resistance mechanism, cultivar selection and help to control the phytoplasma disease.