摘要
槟榔是海南省重要的热带经济作物,由植原体侵染引起的槟榔黄化病(areca palm yellow leaf disease,YLD)是当前我国槟榔生产上的一种毁灭性病害。为了建立精准高效的槟榔黄化植原体检测方法,本研究基于槟榔黄化植原体16SrDNA基因,设计并合成特异性引物AMf/AMr和探针AM-Prode,使用该方法进行准确性、敏感性、特异性及重复性测试,并在其他植物的植原体病害进行检测。结果显示:本检测方法能够准确的检测出阳性样品,健康样品无扩增曲线;在敏感性测试中,该检测方法能检测到1.16×10^(1) copies/μL样本浓度水平,其标准曲线方程为y=-3.4185x+43.624,扩增效率为96.12%,相关系数R^(2)=0.9833;在特异性测试中,该检测方法对YLD的检测具有较好的特异性,槟榔、槟榔其他病害病原及其内生菌的基因组对本方法未造成干扰;在重复性测试中,该检测方法对槟榔黄化病的检测具有较好的重复性;并且该检测方法可对苦楝黄化病、细圆藤丛枝病及辣椒黄化病等8种植原体病害进行检测,对植原体的检测具有一定的通用性。本检测方法的建立,有利于为槟榔黄化病的精准诊断、病原监测及媒介昆虫的检测等研究提供可靠的技术手段。
Areca palm is an important tropical economic crop in Hainan Province.Areca palm yellow leaf disease(YLD)caused by phytoplasma infection is a devastating disease in the production of areca in China.In order to establish an accurate and efficient detection method for areca palm yellow leaf phytoplasma,this study designed and synthesized specific primers AMf/AMr and AM-Prode based on the 16S rDNA gene of areca palm yellow leaf phytoplasma.The method was used to test the accuracy,sensitivity,specificity and repeatability,and to detect phytoplasma diseases in other plants.The method could accurately detect the positive samples,and the healthy samples had no amplification curve.In the sensitivity test,the method could detect the sample concentration level of 1.16×10^(1) copies/μL,and the standard curve equation was y=-3.4185x+43.624,the amplification efficiency was 96.12%,and the correlation coefficient R^(2)=0.9833.In the specificity test,the method had good specificity for the detection of YLD,and the genomes of areca,other disease pathogens and the endophytes did not interfere with the method.In the repeatability test,the method had good repeatability for the detection of YLD.The detection method could be used to detect 8 phytoplasma diseases such as chinaberry yellow leaf disease,‘Pericampylus glaucus’witches'broom disease and pepper yellow leaf disease and the detection of phytoplasma has certain universality.The establishment of detection method is conducive to providing reliable technical means for the accurate diagnosis of YLD,pathogen monitoring and vector insect detection.
作者
林兆威
孟秀利
唐庆华
牛晓庆
宋薇薇
LIN Zhaowei;MENG Xiuli;TANG Qinghua;NIU Xiaoqing;SONG Weiwei(Coconut Research Institute,Chinese Academy of Tropical Agricultural Sciences/Hainan Innovation Center of Academician Team,Wenchang,Hainan 571339,China)
出处
《热带作物学报》
CSCD
北大核心
2024年第6期1120-1126,共7页
Chinese Journal of Tropical Crops
基金
海南省自然科学基金项目(No.321QN345)
海南省院士创新平台科研专项(No.YSPTZX202138,No.YSPTZX202151)。
