李承达 张亚鑫 刘权德 唐玉琴

摘    要：为建立一种由长枝木霉引发灵芝绿霉病的快速检测体系，通过对形态特征及致病性的测定确定病原菌菌株，针对病原菌的特异性基因设计合成引物，对引物进行特异性和灵敏度试验，再对接种过病原菌的土壤进行灵敏度验证。结果表明，病原菌为长枝木霉，设计的引物具有较高的特异性，检测浓度下限为10-2 ng·μL-1;可从含有4.28×106个孢子的1 g土壤中扩增出条带。相比较于传统的检测方法耗时更短，结果检测性高，具有高特异性和灵敏度。

关键词：长枝木霉;灵芝;快速检测

中图分类号：S435.673         文献标识码：A          DOI 编码：10.3969/j.issn.1006-6500.2021.07.014

Abstract： In order to establish a rapid detection system of green mold on Ganoderma lingzhi caused by Trichoderma longibrachiatum，the identification of the pathogen strains was based on determining of morphological and pathogenic characteristics. The study designed and synthesized primers aiming at the specific genes of pathogens， and then carried out specificity and sensitivity tests on the primers. The soil inoculated pathogens were also tested. The results indicated that the pathogen was Trichoderma longibrachiatum. The designed primers have high specificity and the minimum testing concentration was 10-2 ng·μL-1. In the polymerase chain reaction （PCR） for Genomic DNA of soil inoculated pathogens， products occurred in the 1 g soil contained 4.28×106 spores with the designed primers. Compared with the traditional detection methods， the system is more accurate and costs less time.

Key words： Trichoderma longibrachiatum; Ganoderma lingzhi; rapid detection

灵芝（Ganoderma lingzhi）属担子菌门伞菌纲多孔菌目灵芝科[1]。灵芝是我国传统名贵药材，具有多种生理活性和药理作用[2]。中国人工栽培灵芝已有60多年历史， 主要分布在四川、山东、广东、广西、福建、江西、浙江及吉林等地，栽培模式主要以覆土栽培为主[3]。近年来，灵芝市场需求大约每年在以18%～30%的速度增长[4]，2015年我国灵芝栽培面积已达到1万hm2， 灵芝及孢子粉产量约12万t， 产值16亿美元，分别约占全球的75%和30%，已成为世界上灵芝的主要生产和出口国[5]。随着灵芝市场的走俏，其栽培从业者与日俱增，随之发生的病害也作为一个重要的研究方向来开展。灵芝菌丝体生长初期颜色洁白、浓密，绿霉染病初期菌丝体为灰白色、致密，因此早期病害特征肉眼难以辨别。当菌椴颜色逐渐变为浅绿直至深绿色时，轻者导致灵芝减产，重者则绝收。且连作过程中土壤积累的有害微生物导致灵芝老产区绿霉菌源较多，造成灵芝子实体生长缓慢、子实体变小、畸形等现象[6]。……