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为了探索活性内生真菌ZPRa-R-1对宿主DNA遗传性状是否存在影响,采用ISSR分子标记法对二者互作15d的大花红景天DNA多态性进行分析研究。结果表明,采用12条ISSR引物对ZPRa-R-1接种的红景天进行扩增,可扩增出94条条带,其中多态性条带为82条,其百分比为87.23%。每条引物平均扩增7.83条带,扩增片段长度在100~8 000bp之间;菌苗互作15d与对照组的Nei’s遗传距离和遗传一致度分别为0.123 3~1.285 2和0.276 6~0.884 0;真菌接种的大花红景天、无菌组培苗和真菌ZPRs-R-11之间的基因流Nm=0.138 8,小于1;聚类分析得到三个遗传性状分支,第Ⅰ大类包括对照组无菌大花红景天及真菌与组培苗共培养1~10d的样品;第Ⅱ大类为真菌与组培苗共培养11~15d的样品;第Ⅲ大类为对照组ZPRa-R-1真菌;在接种的15d内,共生时间越长,二者之间的遗传相似系数越大,表明活性内生真菌ZPRa-R-1能够引起宿主红景天遗传性状改变。
In order to explore whether ZPRa-R-1, an active endophytic fungus, affects DNA genetic traits, DNA polymorphism of Rhodiola rosea was analyzed by ISSR molecular marker method. The results showed that 12 ISSR primers were used to amplify ZPRa-R-1 inoculated Rhodiola rosea, 94 bands were amplified, of which 82 bands were polymorphic, with a percentage of 87.23%. The average length of each amplified fragment was 7.83 and the length of amplified fragment ranged from 100 to 8 000 bp. The Nei’s genetic distance and genetic identity between the two groups were 0.123 3 ~ 1.285 2 and 0.276 6 ~ 0.884 0; the gene flow Nm = 0.138 8, less than 1 between the fungal inoculated Rhodiola rosea, the sterile tissue culture plant and the fungus ZPRs-R-11; the cluster analysis obtained three branches of genetic traits, The control group of sterile Rhodiola rosea and fungi and tissue culture seedlings co-cultured 1 ~ 10d samples; the second category is fungi and tissue culture seedlings co-cultured for 11 ~ 15d samples; the third group for the control group ZPRa-R -1 fungi. The longer the duration of the symbiosis, the greater the genetic similarity between the two cultivars during the 15 days of inoculation, indicating that the active endophyte ZPRa-R-1 can cause the genetic variation of the host Rhodiola rosea.