荆条种子无菌萌发过程中污染菌的分离鉴定及抑菌剂筛选  

Isolation,Identification of Contaminating Fungi and Screening of Fungistatic Agents in Sterile Germination Process of Seeds of Vitex negundo

作　　者：李宇霞 王霞 张敏[1] 邱悦 张袁菲 梁慧 梁丽琴 陈惠 LI Yuxia;WANG Xia;ZHANG Min;QIU Yue;ZHANG Yuanfei;LIANG Hui;LIANG Liqin;CHEN Hui(Modern College of Humanities and Sciences,Shanxi Normal University,Linfen 041000,China;School of Life Science,Shanxi Normal University,Taiyuan 030000,China)

机构地区：[1]山西师范大学现代文理学院,山西临汾041000 [2]山西师范大学生命科学学院,山西太原030000

出　　处：《山西农业科学》2023年第3期249-256,共8页Journal of Shanxi Agricultural Sciences

基　　金：山西师范大学现代文理学院基础研究项目(2019JCYJ19,2019JCYJ21)。

摘　　要：荆条具有生态、观赏、药用及实用等方面的价值,但在荆条快速繁殖体系的建立中发现,种子在无菌萌发过程中污染率极高,达50%~75%。为降低荆条种子无菌萌发过程中的污染率,试验通过鉴定其主要污染菌株,筛选出有效抑菌剂,并利用组织分离法对污染菌进行分离和纯化;同时,通过光学显微镜和扫描电镜对污染菌的分生孢子进行细胞学观察,然后采用真菌rDNA基因的ITS序列通用引物进行分子生物学鉴定,最后选择百菌清、代森锰锌、多菌灵、氟吗锰锌4种杀菌剂添加到PDA培养基中进行抑菌效果分析。结果表明,主要污染菌为种子表面附着的真菌,菌落呈绒状黄绿色同心圆形,菌丝为有隔褐色分支状菌,分生孢子为橄榄棕色、有隔、梭型;细胞学和分子生物学鉴定表明,该污染菌为链格孢菌。抑菌研究结果发现,60%氟吗锰锌对链铬孢菌的抑制效果最佳,菌落直径最小。将60%氟吗锰锌稀释1 400倍添加到荆条种子无菌萌发培养基(水琼脂培养基)中,污染率明显下降到5.5%。荆条种子无菌萌发中主要污染菌为链格孢菌,60%氟吗锰锌对其抑制效果最佳。Vitex negundo has ecological, ornamental, medicinal, and practical values. However, in the establishment of the rapid propagation system of Vitex negundo, it was found that the contamination rate of seeds in the process of sterile germination was extremely high, reaching 50%-75%. In this study, in order to reduce its contamination rate in the process of sterile germination of Vitex negundo seeds, by identifying its main contaminating strains, and screening out effective fungistatic agents, tissue separation method was used to isolate and purify the contaminating fungi. Simultaneously, the cytological observation of the conidia of the contaminating fungi was carried out by light microscope and scanning electron microscope. Then the universal primers of the Internal Transcribed Spacer(ITS) sequence of the fungal rDNA gene were used for molecular identification. Finally, four fungicides, including chlorothalonil, mancozeb, carbendazim, and Flumorph-Mn-Zn were added to the PDA medium for analysis of antifungial effect. The results showed that the main contaminating fungi were the fungi located on the surface of the seeds. The colonies were yellowish-green concentric circles in velvety shape. The mycelia were septum brown branching bacteria, and the conidia were olive brown, septum and clostridial form. Cytological and molecular biological identifications indicated that the contaminating fungus was Alternaria alternata. The antifungial study showed that 60% of Flumorph-Mn-Zn had the best inhibitory effect on Alternaria alternata. and the colony diameter was the smallest. The contamination rate was significantly reduced to 5.5%, when 60% of Flumorph-Mn-Zn was diluted 1 400 times and added to the sterile germination medium(water agar medium) of Vitex negundo seeds. In conclusion, the main contaminating fungus in the sterile germination of Vitex negundo seeds was Alternaria altemate, 60% of Flumorph-Mn-Zn had the best inhibitory effect on it.

关 键 词：荆条种子 无菌萌发 污染真菌 链格孢菌 氟吗锰锌 

分 类 号：S793.7[农业科学—林木遗传育种]