Cai C, Yang Y, Cheng L, Tong C, Feng J
Genetika. 2015 Jun;51(6):694-703. doi: 10.7868/s001667581502006x.
Because of the advantages of EST-SSR markers, it has been employed as powerful markers for genetic diversity analysis, comparative mapping and phylogenetic studies. In this study, a total of 429,869 tobacco (Nicotiana tabacum L.) ESTs were downloaded from the public databases, which offers an opportunity to identify SSRs in ESTs by data mining, and 38,165 SSRs were identified from 379,967 uni-ESTs with the frequency of one SSR per 5.52 kb. Mono- and tri-nucleotide repeat motifs were the dominant repeat types, accounting for 40.53 and 34.51% of all SSRs, respectively. After eliminating mononucleotide-containing sequences, 86 pairs of primers were designed to amplify in four tobacco accessions. Only 15 primers (17.44%) showed polymorphism, and then they were further used to assess genetic diversity of 20 tobacco accessions. Unweighted pair-group method with arithmetic average dendrograms (UPGMA) and principal coordinates analysis plots (PCA) revealed genetic differentiation between N. rustica and N. tabacum, and between oriental tobacco and other accessions of N. tabacum. The present study reported the development of EST-SSR markers in tobacco by exploiting EST databases, and confirmed the effective way to develop markers. These EST-SSRs can serve in studies on cultivar identification, genetic diversity analysis, and genetics in tobacco.
由于EST-SSR标记的优点，它已被用作遗传多样性分析、比较作图和系统发育研究的有力标记。在本研究中，从公共数据库下载了总共429,869条烟草（Nicotiana tabacum L.）EST序列，这为通过数据挖掘在EST中鉴定SSR提供了机会，并且从379,967条单基因EST中鉴定出38,165个SSR，频率为每5.52 kb一个SSR。单核苷酸和三核苷酸重复基序是主要的重复类型，分别占所有SSR的40.53%和34.51%。去除含单核苷酸的序列后，设计了86对引物在4个烟草种质中进行扩增。只有15对引物（17.44%）表现出多态性，然后将它们进一步用于评估20个烟草种质的遗传多样性。非加权算术平均法聚类图（UPGMA）和主坐标分析图（PCA）揭示了黄花烟草与普通烟草之间以及香料烟与普通烟草的其他种质之间的遗传分化。本研究报道了通过利用EST数据库开发烟草EST-SSR标记，并证实了开发标记的有效方法。这些EST-SSR可用于烟草品种鉴定、遗传多样性分析和遗传学研究。
