一种TMV抗性离体快速鉴定方法及其在烟草育种上的应用  

A Rapid Identification Method for TMV Resistance in Vitro and Its Application in Tobacco Breeding

作　　者：殷宁宁 余文 巫升鑫 程崖芝 赖聚贤 陈顺辉 吴为人 高文霞 Yin Ningning;Yu Wen;Wu Shengxin;Cheng Yazhi;Lai Juxian;Chen Shunhui;Wu Weiren;Gao Wenxia(Fujian Key Laboratory of Crop Breeding by Design,College of Agriculture,Fujian Agriculture and Forestry University,Fuzhou,350002;Institute of Tobacco Science,Tobacco Monopoly Administration(Company)of Fujian Province,Fuzhou,350003;Key Laboratory of Genetics,Breeding and Multiple Utilization of Crops,Ministry of Education,College of Agriculture,Fujian Agriculture and Forestry University,Fuzhou,350002;Key Laboratory of Biological Breeding for Fujian and Taiwan Crops(Ministry of Agriculture and Rural Affairs),College of Agriculture,Fujian Agriculture and Forestry University,Fuzhou,350002)

机构地区：[1]福建农林大学农学院,福建省作物设计育种重点实验室,福州350002 [2]福建省烟草专卖局,烟草科学研究所,福州350003 [3]福建农林大学农学院,作物遗传育种与综合利用教育部重点实验室,福州350002 [4]福建农林大学农学院,闽台作物生物育种农业农村部重点实验室,福州350002

出　　处：《分子植物育种》2023年第24期8144-8150,共7页Molecular Plant Breeding

基　　金：中国烟草总公司福建省烟草公司科技计划项目(2019350000240136)资助。

摘　　要：烟草普通花叶病,由烟草花叶病毒(tobacco mosaic virus,TMV)引起,是烟草生产中的常见病害之一,培育和种植抗病品种是解决该病害的主要途径,育种材料的抗性鉴定是抗病育种的关键环节。为提高TMV抗性鉴定效率,本研究根据TMV与N基因作用的原理设计了一种烟草苗期离体接种快速鉴定TMV抗性的方法。该方法为研磨病害严重程度为5级的新鲜病叶(10 g)并过滤,往过滤液中加纯净水(100 m L)稀释为病毒液;摘取6~7叶期烟苗的叶片(从上往下数第3叶)进行离体摩擦接种;3 d后观察坏死斑发生情况。应用此鉴定方法对20份烟草材料的TMV抗性进行鉴定,结果表明13份材料出现免疫斑;同时,对20份烟草材料的N基因进行了检测,结果表明13份材料含有N基因。出现免疫斑的材料与含N基因的材料相一致,证明了苗期离体接种方法鉴定TMV抗性的可行性。将此方法应用于21524份育种中间材料(7047株BC_(1)F_(3)、10013株BC_(1)F_(4)、4464株BC_(1)F_(5))的TMV抗性鉴定中,结果得到9829株(3103株BC_(1)F_(3),4762株BC_(1)F_(4),1964株BC_(1)F_(5))抗性植株。鉴定结果基本符合10761株抗性植株(3523株BC_(1)F_(3),5006株BC_(1)F_(4),2232株BC_(1)F_(5))的理论值,进一步证明了此方法的可靠性。本研究对大量烟草育种中间材料的TMV抗性鉴定具有重要的应用价值。Tobacco common mosaic disease,caused by tobacco mosaic virus (TMV),is one of the common disease in tobacco production.Breeding and cultivating disease-resistant varieties is the main approach to control tobacco common mosaic disease,and the resistance identification of breeding materials is a crucial step in disease-resistant breeding.In order to improve the efficiency of TMV resistance identification,this study designed a method for rapid identification of TMV resistance by in vitro inoculation at tobacco seedlings based on the interaction between TMV and the N gene.The method involved grinding fresh diseased leaves with a severity level of 5 (10 g)and filtering the extract,diluting it with distilled water (100 mL) to obtain a virus solution.The third leaf from the top of 6~7 leaf stage tobacco seedlings was taken for in vitro friction inoculation,and the occurrence of necrotic spots was observed after 3 days.This identification method was applied to evaluate the TMV resistance of 20 tobacco materials,and the results showed that 13 materials exhibited immune spots.Simultaneously,the N gene of the 20 tobacco materials was detected and 13 materials were found to contain the N gene.The consistency between the occurrence of immune spots and the presence of the the N gene confirmed the feasibility of the in vitro inoculation method for TMV resistance identification.This method was further applied to evaluate the TMV resistance of21 524 breeding intermediate materials (7 047 BC_1F_3,10 013 BC_1F_4,4 464 BC_1F_5),and the results showed that9 829 plants (3 103 BC_1F_3,4 762 BC_1F_4,1 964 BC_1F_5) showed resistance.The identification results were basically consistent with the theoretical value of 10 761 resistant plants (3 523 BC_1F_3,5 006 BC_1F_4,2 232 BC_1F_5),which further confirmed the reliability of this method.This study has important application value for TMV resistance identification of a large number of intermediate materials in tobacco breeding.

关 键 词：TMV抗性 N基因 离体接种 快速鉴定方法 育种应用 

分 类 号：S572[农业科学—烟草工业]