Development and Characterization of Genic SSR Markers from Indian Mulberry Transcriptome and Their Transferability to Related Species of Moraceae

Improving mulberry leaf production with enhanced leaf quality holds the key to sustain the ever increasing demand for silk. Adoption of modern genomic approaches for crop improvement is severely constrained by the lack of sufficient molecular markers in mulberry. Here, we report development and validation of 206 EST derived SSR markers using transcriptome data generated from leaf tissue of a drought tolerant mulberry genotype, Dudia white. Analysis of transcriptome data containing 10169 EST sequences, revealed 1469 sequences with microsatellite repeat motifs. We designed a total of 264 primers to the most appropriate repeat regions, of which 206 were locus specific. These markers were validated with 25 diverse mulberry accessions and their transferability to closely related species belonging to family Moraceae was examined. Of these markers, 189 revealed polymorphism with up to 8 allelic forms across mulberry species, genotypes and varieties with a mean of 3.5 alleles per locus. The markers also revealed higher polymorphic information content of 0.824 among the accessions. These markers effectively segregated the species and genotypes and hence, can be used for both diversity analysis and in breeding applications. Around 40% of these markers were transferable to other closely related species. Along with the other genic and genomic markers, we report a set of over 750 co-dominant markers. Using these markers we constructed the first genetic linkage map of mulberry exclusively with co-dominant markers.

提升桑叶产量与叶片品质，是满足持续增长的蚕丝需求的核心所在。当前，现代基因组学技术在作物改良中的应用，因桑属植物缺乏足够的分子标记而受到严重限制。本研究利用耐旱桑基因型“杜迪白（Dudia white）”的叶片组织转录组数据，开发并验证了206个表达序列标签-简单序列重复（Expressed Sequence Tag-Simple Sequence Repeat, EST-SSR）标记。对包含10169条表达序列标签（Expressed Sequence Tag, EST）序列的转录组数据进行分析后，共筛选得到1469条含有微卫星重复基序的序列。针对其中最适宜的重复区域，我们共设计了264对引物，其中206对具有位点特异性（locus specific）。我们利用25份不同的桑属种质对这些标记进行了验证，并检测了其对桑科（Moraceae）近缘物种的跨物种可转移性。结果显示，189个标记在不同桑属物种、基因型及品种间呈现多态性，每一位点的等位基因数最多可达8个，平均每一位点含3.5个等位基因。该类标记在供试种质间的多态性信息含量（polymorphic information content, PIC）均值高达0.824。这些标记可有效区分不同物种与基因型，因此可同时用于种质多样性分析与育种应用。其中约40%的标记可在其他近缘物种中实现跨物种扩增。结合已有的其他基因标记与基因组标记，本研究共报道了超过750个共显性标记（co-dominant markers）。利用上述标记，我们构建了首张仅使用共显性标记的桑属植物遗传连锁图谱（genetic linkage map）。