王泽平 宋修鹏 颜梅新 李毅杰 李翔 张小秋 唐红琴 龙盛风

摘要 本文旨在探討核苷酸结合位点和富含亮氨酸重复（NBS-LRR）类基因参与甘蔗抗梢腐病菌侵染应答机制，为后续克隆关键抗病基因以及研究抗病机理提供理论依据。试验利用Illumina高通量转录组测序技术检测高抗梢腐病品种‘粤糖94-128和高感梢腐病品种‘桂糖37号接种前后NBS-LRR类抗梢腐病基因的表达情况，然后设计引物对显著差异表达基因进行不同接种时期下的荧光定量PCR验证。结果表明，16个NBS-LRR类基因在甘蔗叶片受到梢腐病菌侵染后持续上调表达，但表达趋势存在两种情况，13个基因在诱导后7 d显著或极显著上调表达，3个基因在诱导7 d后上调表达不显著，在诱导14 d后才显著上调表达。据此可将其分为瞬时基础防御基因（0～7 d）和滞后特异性防御基因（14～21 d），确定NBS-LRR类基因参与甘蔗防御梢腐病菌的侵染。

关键词 甘蔗; 梢腐病; 转录组; NBS-LRR

中图分类号： S 556.1， S 435.661

文献标识码： A

DOI： 10.16688/j.zwbh.

2020301

Screening and quantitative analysis of NBS-LRR gene against pokkah boeng disease in sugarcane

WANG Zeping1， SONG Xiupeng1， YAN Meixin1， LI Yijie1， LI Xiang1，ZHANG Xiaoqiu1， TANG Hongqin2*， LONG Shengfeng3*

（1. Sugarcane Research Institute， Guangxi Academy of Agricultural Sciences， Sugarcane Research Center，

Chinese Academy of Agricultural Science， Key Laboratory of Sugarcane Biotechnology and Genetic

Improvement （Guangxi）， Ministry of Agriculture and Rural Affairs， Nanning 530007， China;

2. Institute of Agricultural Resources and Environment， Guangxi Academy of Agricultural Sciences，

Nanning 530007， China; 3. Guangxi Academy of Agricultural Sciences， Nanning 530007， China）

Abstract

The molecular mechanism of nucleotide binding sites and leucine-rich replication （NBS-LRR） genes resistant to pokkah boeng disease （PBD） in sugarcane was explored to provide a theoretical basis for cloning resistant genes. This study firstly used Illumina high-throughput transcriptome sequencing technology to detect NBS-LRR genes against PBD with typical resistant and susceptive sugarcane materials， and then designed primers to verify the expression characteristics of significant differentially expressed NBS-LRR genes under different inoculation periods using real-time quantitative PCR （qRT-PCR）. The results showed that 16 NBS-LRR-like genes steadily increased expression when sugarcane leaves were infected with the PBD pathogen， including two trends in expression： 13 genes significantly or extremely significantly increased expression at 7 d post-inoculation， but three genes did not significantly increased expression at 7 d post-inoculation， but significantly increased expression at 14 d post-inoculation. Therefore， these genes could be divided into instantaneous basic defense genes （0 d to 7 d） and specific lagging defense genes （14 d to 21 d）， suggesting that NBS-LRR genes are actively involved in resistance against the PBD pathogen.

Key words

sugarcane; pokkah boeng disease; transcriptome; NBS-LRR

甘蔗梢腐病（pokkah boeng disease）作为一种真菌性气传病害，其发生几乎遍及所有的甘蔗生产国家和地区，在我国主产蔗区广西已上升为仅次于黑穗病的主要病害[1]。生产上主要是通过喷施多菌灵进行防治，而挖掘抗病基因进而利用其进行杂交聚合育种是解决当前主栽甘蔗品种抗病性有限的重要途径。随着克隆技术的发展，目前对植物抗病基因的结构有了全面的认识，在许多植物中已克隆出抗病基因，且多种植物的核苷酸结合位点和富含亮氨酸重复（nucleotide binding sites and leucine-rich replication，NBS-LRR）类抗病基因的数目已确定[2]。……