Gupta P K, Rustgi S, Sharma S, Singh R, Kumar N, Balyan H S
Molecular Biology Laboratory, Department of Agricultural Botany, Ch. Charan Singh University, 250 004 (U.P.), Meerut, India.
Mol Genet Genomics. 2003 Dec;270(4):315-23. doi: 10.1007/s00438-003-0921-4. Epub 2003 Sep 24.
Nearly 900 SSRs (simple sequence repeats) were identified among 15,000 ESTs (expressed sequence tags) belonging to bread wheat ( Triticum aestivumL.). The SSRs were defined by their minimum length, which ranged from 14 to 21 bp. The maximum length ranged from 24 to 87 bp depending upon the length of the repeat unit itself (1-7 bp). The average density of SSRs was one SSR per 9.2 kb of EST sequence screened. The trinucleotide repeats were the most abundant SSRs detected. As a representative sample, 78 primer pairs were designed, which were also used to screen the dbEST entries for Hordeum vulgare and Triticum tauschii (donor of the D-genome of cultivated wheat) using a cut-off E (expectation) value of 0.01. On the basis of in silico analysis, up to 55.12% of the primer pairs exhibited transferability from Triticum to Hordeum, indicating that the sequences flanking the SSRs are not only conserved within a single genus but also between related genera in Poaceae. Primer pairs for the 78 SSRs were synthesized and used successfully for the study of (1) their transferability to 18 related wild species and five cereal species (barley, oat, rye, rice and maize); and (2) polymorphism between the parents of four mapping populations available with us. A subset of 20 EST-SSR primers was also used to assess genetic diversity in a collection of 52 elite exotic wheat genotypes. This was done with a view to compare their utility relative to other molecular markers (gSSRs, AFLPs, and SAMPL) previously used by us for the same purpose with the same set of 52 bread wheat genotypes. Although only a low level of polymorphism was detected, relative to that observed with genomic SSRs, the study suggested that EST-SSRs can be successfully used for a variety of purposes, and may actually prove superior to SSR markers extracted from genomic libraries for diversity estimation and transferability.
在属于普通小麦（Triticum aestivum L.）的15000个表达序列标签（EST）中鉴定出了近900个简单序列重复（SSR）。这些SSR根据其最短长度来定义，最短长度范围为14至21 bp。最长长度范围为24至87 bp，这取决于重复单元本身的长度（1至7 bp）。筛选的EST序列中SSR的平均密度为每9.2 kb有一个SSR。三核苷酸重复是检测到的最丰富的SSR。作为代表性样本，设计了78对引物，这些引物还用于筛选大麦（Hordeum vulgare）和节节麦（Triticum tauschii，栽培小麦D基因组的供体）的dbEST条目，使用的截止期望（E）值为0.01。基于电子分析，高达55.12%的引物对表现出从小麦到大麦的可转移性，这表明SSR侧翼序列不仅在单个属内保守，而且在禾本科的相关属之间也保守。合成了78个SSR的引物对，并成功用于以下研究：（1）它们向18个相关野生种和5个谷物种（大麦、燕麦、黑麦、水稻和玉米）的可转移性；以及（2）我们现有的四个作图群体亲本之间的多态性。还使用了20个EST-SSR引物的子集来评估52个优良外来小麦基因型群体的遗传多样性。这样做是为了将它们的效用与我们之前用于同一目的、针对同一组52个普通小麦基因型使用的其他分子标记（基因组SSR、AFLP和SAMPL）进行比较。尽管相对于基因组SSR观察到的多态性水平较低，但该研究表明EST-SSR可成功用于多种目的，并且实际上可能在多样性估计和可转移性方面优于从基因组文库中提取的SSR标记。
