Identification and functional annotation of expressed sequencetags based SSR markers of Stevia rebaudiana

Authors: RAJINDER KAUR, NEHA SHARMA, RAVINDER RAINA

Abstract: Stevia rebaudiana Bertoni, a perennial herb belonging to the family Asteraceae, is a tropical as well as subtropical plant of medicinal importance. Despite being an important medicinal plant and in fact proving a boon to diabetics, it has remained almost ignored by molecular biologists. A systematic molecular evaluation of stevia germplasm and molecular breeding work are sparsely reported mainly due to the limited specific molecular markers available. Expressed sequence tags (ESTs) are a valuable resource for developing simple sequence repeats (SSRs). In this study, 5548 stevia ESTs sequences from leaf tissues were retrieved from the NCBI database. Clustering and assembly of these ESTs resulted in a nonredundant set of sequences, i.e. A total of 471 contigs and 3845 singleton ESTs. Out of these 471 contigs and 3845 singletons, 168 SSRs were identified. Dinucleotide SSR is the dominant repeat type (62.5%), followed by trinucleotide (37.5%). Annotation analysis revealed that putative function could be assigned to 82.2% of EST-SSRs. After using Primer3 software, 18 primers were custom synthesized from the SSR containing 18 singletons and generated 61.11% polymorphism. In silico mined markers with functional annotations, in the present study, facilitated genome analysis in stevia, which had not been performed previously. Additionally, the EST-SSRs will be used for molecular work in related plant species since they generally exhibit cross species transferability, making further work cost effective and simple.

Keywords: EST, functional annotation, genetic diversity, SSR, Stevia rebaudiana

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