菊花EST-SSR标记的开发与应用

菊花EST-SSR标记的开发与应用 Development and Application of EST-SSR Markers in Garden Chrysanthemum（Chrysanthemum grandiflora Ramet.） 张运兴 ZHANG Yunxing 1 first-author 李卫国 LI Weiguo 1 申亚琳 SHEN Yalin 1 杨伟鑫 YANG Weixin 1 河南理工大学资源环境学院 河南理工大学资源环境学院 Institute of Resources and Environment,Henan Polytechnic University Institute of Resources and Environment,Henan Polytechnic University 从NCBI（National Center for Biotechnology Information）数据库下载7 259条菊花相关表达序列标签（EST）序列,经去除冗余序列拼接后得到3 784条Uni-ESTs,总长度为2 088.6kb.用MISA软件查找其简单重复序列（SSR）位点,共获得407个SSR位点,分布于355条EST上,出现的频率为10.76%.在搜索到的EST-SSR中,三核苷酸重复类型占主导地位,所占比例为47.42%.共观察到76种重复基元,出现最多的重复基元是A/T和ACC/GGT,其次为ATC/ATG,AAC/GTT,AC/GT,AAT/ATT.根据搜索结果随机设计了24对菊花EST-SSR引物,对14个菊花品种进行PCR扩增,发现有12对引物有较清晰且稳定的扩增产物（其中10对引物的产物存在多态性）,共检出37个位点（其中多态性位点32个）,平均每对引物可扩增出3.2条多态性片段.遗传相似性分析显示,14个品种的遗传相似系数在0.61～0.84之间,平均相似系数为0.67. A total of 7 259 garden Chrysanthemum expression sequence tags（EST） were downloaded from NCBI（National Center for Biotechnology Information） databases,and were further assembled to 3 784 non-redundant ESTs with a total length of 2 088.6 kb.Of the assembled sequences,407 SSRs were identified in 355 Uni-ESTs using MISA software,and the frequency of the EST-SSR was 10.76%.Trinucleotide repeats were major types,and accounted for 47.42% of the total SSR.Among the 76 observed repetitive motifs,A/T and ACC/GGT were the most abundant motifs,followed by ATC/ATG,AAC/GTT,AC/GT,and AAT/ATT.Based on the information of EST-SSR in Chrysanthemum,24 EST-SSR primer pairs were randomly selected to assess the amplification efficiency by using 14 Chrysanthemum cultivars,and 12 primer pairs yielded ideal PCR products.A total of 37 loci,which includes 32 polymorphic loci were amplified by using 10 SSR primer pairs,ranging from 3-5 loci per primer pair with an average of 3.2.Among 14 Chrysanthemum cultivars,genetic similarity ranged from 0.61 to 0.84 with a mean of 0.67. 菊花 表达序列标签（EST） 简单重复序列（SSR） 遗传多样性 garden Chrysanthemum express sequence tag simple sequence repeat genetic diversity 10.14188/j.1671-8836.2013.04.011 S682.11 国家自然科学基金（31100182）;河南理工大学博士基金（B2008-25）资助项目 2013-04-01 2021-04-01 4 59