不同茯苓菌株的质量评价及遗传关系鉴定

Quality evaluation and genetic relation identification for different Poria cocos strains

【目的】评价不同茯苓菌株质量并鉴定其遗传关系,为茯苓菌株资源合理利用及优良菌株选育提供参考。【方法】以11株不同来源的茯苓菌株为研究对象,分别采用平面培养法测定其菌丝生长速率,松树兜栽培法测定其菌核产量,反复转接法测定其遗传稳定性。提取不同茯苓菌株的总DNA,PCR扩增ITS序列,进行ITS序列分析,并构建系统发育进化树。同时检测菌株间的菌丝拮抗性,结合ITS序列分析结果进一步鉴定茯苓菌株遗传关系。【结果】11个供试菌株中,8、9和12号菌株的菌丝生长速率、菌核产量和遗传稳定性均较高,1、7和13号菌株较低。茯苓ITS序列长度为1600 bp,其序列分析结果显示,1、2、3、4、7和10号菌株与Wolfporia cocos intemal transcribed spacer 1的同源性高达100%,8、9、11、12和13号菌株与W.cocos strain LP-13的同源性达99%,表明供试茯苓菌株属于两个不同的亚种。系统发育进化树分析结果显示,1、2、3、4、7和10号菌株聚为一类,菌株间的遗传关系非常接近,存在同种异名的可能;8、9、11、12和13号菌株聚为另一类,菌株间的遗传距离较大,存在种内差异。【结论】通过测定菌丝生长速度、菌核产量和遗传稳定性评价茯苓菌株质量切实可行,ITS序列分析可有效将茯苓菌株鉴定到种级分类单元,并明确茯苓菌株间的遗传关系。

【Objective】Qualities of different Poria cocos strains were evaluated in the study and their genetic relations were identified to provide reference for the rational utilization of P.cocos and breeding of fine strains.【Method】Eleven P.cocos strains from various sources were research objects.Their mycelial growth rates were detected through planar culture,sclerotium yields were determinated through cultivation using pine tree pocket,genetic stabilities were identified through repeated switching method.The genomes were extracted from these strains,ITS sequence were amplified by PCR and then ITS sequences were analyzed,and phylogenetic tree was established.Meanwhile,hypha antagonism among the strains was detected,and the genetic relations of P.cocos strains were further investigated combining ITS sequence results.【Result】Among the 11 tested strains,mycelial growth rate,sclerotium yield and genetic stability of strains No.8,No.9 and No.12 were better than others,but those of strains No.1,No.7 and No.13 ranked the bottom.ITS sequence length of P.cocos was 1600 bp.According to the sequence analysis,homology of strains No.1,No.2,No.3,No.4,No.7 and No.10 was 100%with Wolfporia cocos intemaltranscribed spacer1,and the homology of strains No.8,No.9,No.11,No.12 and No.13 was 99%with Wolfporia cocos strain LP-13.These results indicated that the tested strains belonged to two subspecies.Phylogenetic tree indicated that,strains No.1,No.2,No.3,No.4,No.7 and No.10 clustered into one group,and their genetic relations were close,which meant these strains might be the same species with different names.Strains No.8,No.9,No.11,No.12 and No.13 clustered into a group with large genetic relations among them,which indicated that there might be difference within the species.【Conclusion】Evaluating the quality of P.cocos via determinating mycelial growth rate,sclerotium yield and hereditary stability is viable.ITS sequence analysis can identify the strains to species level,and explicit the genetic relationship between strains.