基于ISSR的野生桑树桑黄菌株遗传多样性分析
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摘要
为保护开发野生桑树桑黄种质资源,对17个不同来源的野生桑树桑黄菌株开展种内遗传多样性分析,利用简单重复序列间扩增(ISSR)分子标记技术对桑黄菌株DNA进行扩增,分析扩增条带,利用NTSYS软件构建亲缘关系UPGMA聚类图。结果表明:16条ISSR引物中,有10条ISSR引物多态性丰富,条带清晰;10条引物共检测到904个位点,其中,多态性位点717个,多态性百分比79.3%。在DNA指纹图谱中,引物P5、P812扩增条带多态性最高。NTSYS-PC2.10e软件分析表明,17个桑树桑黄遗传相似系数为0.57～0.99。UPGMA聚类分析结果显示,在遗传相似系数(GS)约0.65处,可将17个桑树桑黄划分为2大类群:S4,S23,S26为一大类群,其余为一大类群。综上可知,桑树桑黄种质资源具有丰富的遗传多样性,ISSR分子标记可有效区分不同桑树桑黄菌株。
In order to provide basis for protection and development of germplasm resources of wild Sanghuangporus sanghuang, the genetic diversity of 17 wild Sanghuangporus sanghuang strains was investigated in this study. The amplified products of 17 wild Sanghuangporus sanghuang strains DNA based on ISSR makers technique were used to analyze the genetic diversity of Sanghuangporus sanghuang. Cluster dendrogram of different samples were established based on the unweighted pair-group method with arithmetic mean(UPGMA) by NTSYS-pc software. The results showed that bands amplified by 10 ISSR primers among the 16 ISSR primers were clear and polymorphisms rich. 10 ISSR primers generated 904 loci, of which 717 loci were polymorphic. The percentage of polymorphisms was 79.3%. Bands amplified by ISSR-primers P5 and P812 had the highest polymorphisms in DNA fingerprint. The genetic similarity coefficients of 17 Sanghuangporus sanghuang strains ranged from 0.57 to 0.99. UPGMA cluster analysis showed that 17 Sanghuangporus sanghuang strains were divided into 2 groups at the level of genetic similarity coefficient(GS) about 0.65 in the cluster dendrogram. S4, S23, S26 were grouped together and the other 14 strains were in one group. Results of ISSR analysis revealed that germplasm resources of Sanghuangporus sanghuang had plentiful genetic diversity. ISSR method was efficient for distinguishing different strains of Sanghuangporus sanghuang.
In order to provide basis for protection and development of germplasm resources of wild Sanghuangporus sanghuang, the genetic diversity of 17 wild Sanghuangporus sanghuang strains was investigated in this study. The amplified products of 17 wild Sanghuangporus sanghuang strains DNA based on ISSR makers technique were used to analyze the genetic diversity of Sanghuangporus sanghuang. Cluster dendrogram of different samples were established based on the unweighted pair-group method with arithmetic mean(UPGMA) by NTSYS-pc software. The results showed that bands amplified by 10 ISSR primers among the 16 ISSR primers were clear and polymorphisms rich. 10 ISSR primers generated 904 loci, of which 717 loci were polymorphic. The percentage of polymorphisms was 79.3%. Bands amplified by ISSR-primers P5 and P812 had the highest polymorphisms in DNA fingerprint. The genetic similarity coefficients of 17 Sanghuangporus sanghuang strains ranged from 0.57 to 0.99. UPGMA cluster analysis showed that 17 Sanghuangporus sanghuang strains were divided into 2 groups at the level of genetic similarity coefficient(GS) about 0.65 in the cluster dendrogram. S4, S23, S26 were grouped together and the other 14 strains were in one group. Results of ISSR analysis revealed that germplasm resources of Sanghuangporus sanghuang had plentiful genetic diversity. ISSR method was efficient for distinguishing different strains of Sanghuangporus sanghuang.
引文
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[10] 秦莲花, 宋春艳, 谭琦, 等. 用ITS和ISSR分子标记技术鉴别香菇生产用种[J]. 菌物学报, 2006, 25(1):94-100. QIN L H, SONG C Y, TAN Q, et al. Use of ITS and ISSR markers to identify cultivated strains of Lentinus edodes[J]. Mycosystema, 2006, 25(1):94-100. (in Chinese with English abstract)
[2] 郎明紫, 曾鹏, 刘明明, 等. 桑黄多糖的研究进展[J]. 蚕桑通报, 2017, 48(2): 14-18.LANG M Z, ZENG P, LIU M M, et al. Research progress on polysaccharide from Phellinus baumii[J]. Bulletin of Sericulture, 2017, 48(2): 14-18. (in Chinese with English abstract)
[3] 邵晨霞, 靳磊, 唐少军, 等. 7株桑黄遗传亲缘关系和栽培特性研究[J].中国食用菌, 2017, 36(3): 40-43.SHAO C X, JIN L, TANG S J, et al. Study on genetic relationships and cultivate characteristic of 7 Inonotus sanghuang strains[J].Edible Fungi of China, 2017, 36(3): 40-43.(in Chinese with English abstract)
[4] 时圣明, 番明佳, 王洁, 等. 分子鉴定技术在中药中的应用[J]. 中草药, 2016, 47(17): 3121-3126.SHI S M, PAN M J, WANG J, et al. Application of molecular identification techniques in Chinese materia medica[J].Chinese Traditional and Herbal Drugs, 2016, 47(17): 3121-3126.(in Chinese with English abstract)
[5] 任梦云, 陈彦君, 张盾, 等. ISSR标记技术在药用植物资源中的研究进展及应用[J]. 生物技术通报, 2017, 33(4): 63-69.REN M Y, CHEN Y J, ZHANG D, et al. Application and research progress of inter-simple sequence repeat (ISSR) marker in medicinal plants[J]. Biotechnology Bulletin, 2017, 33(4): 63-69.(in Chinese with English abstract)
[6] GóESNETO A, LOGUERCIOLEITE C, GUERRERO R T. DNA extraction from frozen field-collected and dehydrated herbarium fungal basidiomata: performance of SDS and CTAB-based methods[J]. Biotemas, 2005 (2): 19-32.
[7] WOLFE A D, XIANG Q Y, KEPHART S R. Diploid hybrid speciation in Penstemon (Scrophulariaceae)[J]. Proceedings of the National Academy of Sciences, 1998, 95(9):5112-5115.
[8] 董彩虹, 刘奇正, 张娇娇. 近十年中国重要食药用菌研究进展[J]. 微生物学杂志, 2017, 37(4): 1-9.DONG C H, LIU Q Z, ZHANG J J. Research progress on important edible and medicinal fungi in China over the last decade[J]. Journal of Microbiology, 2017, 37(4): 1-9.(in Chinese with English abstract)
[9] 任海霞, 宫志远, 蒋志琴, 等. 26个香菇菌株的遗传多样性分析[J]. 山东农业科学, 2017, 49(11):20-23, 44.REN H X, GONG Z Y, JIANG Z Q, et al. Analysis of genetic diversity of 26 Lentinus edodes strains[J]. Shandong Agricultural Sciences, 2017, 49(11):20-23, 44.(in Chinese with English abstract)
[10] 秦莲花, 宋春艳, 谭琦, 等. 用ITS和ISSR分子标记技术鉴别香菇生产用种[J]. 菌物学报, 2006, 25(1):94-100. QIN L H, SONG C Y, TAN Q, et al. Use of ITS and ISSR markers to identify cultivated strains of Lentinus edodes[J]. Mycosystema, 2006, 25(1):94-100. (in Chinese with English abstract)