摘要
【目的】比较基于rDNA序列分析的3种丛枝菌根(AM)真菌分子鉴定方法,为提高AM真菌在种水平分子鉴定的准确性提供参考。【方法】对从柑橘和甘蔗等广西地区主要作物的根际土壤中分离出的AM真菌菌株(编号为A6、B3和GY3-1)进行形态学鉴定,再通过巢式PCR分别扩增AM真菌的18S rDNA、28S rDNA及含ITS1、5.8S rDNA和ITS2的序列(简写为ITS1+5.8S+ITS2),分别将其测序结果与GenBank数据库进行比对,并构建系统发育进化树,再结合形态学鉴定结果比较基于3个rDNA序列分析分子鉴定方法的准确性。【结果】菌株A6、B3和GY3-1的形态特征分别与幼套近明球囊霉(Claroideoglomus etunicatum)、黏质隔球囊霉(Septoglomus viscosum)和摩西斗管囊霉(Funnelifor-mis mosseae)一致。基于18S rDNA序列,将菌株A6鉴定为近明球囊霉属(Claroideoglomus),菌株GY3-1鉴定为摩西斗管囊霉(F.moessae),菌株B3鉴定为黏质隔球囊霉(S.viscosum)。基于28S rDNA序列,将菌株A6鉴定为幼套近明球囊霉(C.etunicatum),菌株B3鉴定为黏质隔球囊霉(S.viscosum),菌株GY3-1鉴定为摩西斗管囊霉(F.moessae)。基于ITS1+5.8S+ITS2序列,将菌株A6鉴定为幼套近明球囊霉(C.etunicatum),菌株B3鉴定为黏质隔球囊霉(S.visco-sum),菌株GY3-1鉴定为摩西斗管囊霉(F.moessae)。而综合3种分子鉴定结果,菌株A6、B3和GY3-1分别鉴定为幼套近明球囊霉(C.etunicatum)、黏质隔球囊霉(S.viscosum)和摩西斗管囊霉(F.mosseae)。【结论】3种方式均可用于AM真菌的在种水平上的分子鉴定,以18S rDNA与28S rDNA为主的鉴定方法相对简单且快速,更适用于对AM真菌在属水平的鉴定;以ITS1+5.8S+ITS2为主的鉴定方法步骤较繁琐,但鉴定至种水平的准确性较高。3种方法的分析结果在一定程度上可互相补充,加强对AM真菌分子鉴定结果的准确性。
【Objective】The purpose of the study was to compare the three molecular identification methods of arbuscu-lar mycorrhizal(AM)fungi based on rDNA sequence analysis,and to provide a reference for improving the accuracy of molecular identification of AM fungi at the species level.【Method】Morphological identification of AM fungi(numbered A6,B3 and GY3-1)isolated from the rhizosphere soil of common crops such as citrus and sugarcane in Guangxi was car-ried out.The 18S rDNA,28S rDNA and sequences containing ITS1,5.8S rDNA and ITS2(abbreviated as ITS1+5.8S+ITS2)of AM fungi were amplified by nested PCR respectively.The sequences obtained by sequencing were respectively aligned with GenBank data and a phylogenetic tree was constructed.The accuracy of the three molecular identification methods based on rDNA sequence analysis was then compared with the morphological identification results.【Result】Mor-phological charaters of A6,B3 and GY3-1 were identical with Claroideoglomus etunicatum,Septoglomus viscosum and Funneliformis mosseae respectively.Based on 18S rDNA sequence,strain A6 was identified as Claroideoglomus,strain GY3-1 as F.moessae,and strain B3 as S.viscosum.Based on 28S rDNA sequence,strain A6 was identified as C.etunica-tum,strain B3 as S.viscosum,and strain GY3-1 as F.moessae.Based on the ITS1+5.8S+ITS2 sequence,strain A6 was identified as C.etunicatum,strain B3 as S.viscosum,and strain GY3-1 as F.moessae.Combining the results of the three molecular identifications,the identification results of A6,B3 and GY3-1 strains were C.etunicatum,S.viscosum and F.mosseae respectively.【Conclusion】All three methods can be used for the molecular identification of AM fungi at the spe-cies level.The identification methods mainly based on 18S rDNA and 28S rDNA are relatively simple and fast,and are more suitable for the identification of AM fungi at the genus level.The identification method mainly based on ITS has more complicated steps,but the accuracy of identification to the species level is higher.The analysis
作者
农颖杰
卢昱帆
许诗萍
高日芳
宋娟
张金莲
韦翔华
陈廷速
NONG Ying-jie;LU Yu-fan;XU Shi-ping;GAO Ri-fang;SONG Juan;ZHANG Jin-lian;WEI Xiang-hua;CHEN Ting-su(College of Agriculture,Guangxi University,Nanning,Guangxi 530004,China;Institute of Microbiology,Guangxi Academy of Agricultural Sciences,Nanning,Guangxi 530007,China;Research Institute of Resources and Plants,Yunnan University,Kunming,Yunnan 650504,China;School of Forestry,Central South University of Forestry and Technology,Changsha,Hunan 410004,China;School of Life Sciences,Jiangxi Normal University,Nanchang,Jiangxi 330022,China)
出处
《南方农业学报》
CAS
CSCD
北大核心
2023年第10期2854-2864,共11页
Journal of Southern Agriculture
基金
广西自然科学基金重点项目(2019GXNSFDA245013)
广西科技基地和人才专项(桂科AD20159001)
广西农业科学院基本科研业务专项(桂农科2023YM96)。
关键词
丛枝菌根真菌
RDNA
系统发育进行树
分子鉴定
arbuscular mycorrhizal fungi
rDNA
phylogenetic tree
molecular identification
