Development of a EST dataset and characterization of EST-SSRs in a traditional Chinese medicinal plant, Epimedium sagittatum (Sieb. Et Zucc.) Maxim

Abstract

BackgroundEpimedium sagittatum (Sieb. Et Zucc.) Maxim, a traditional Chinese medicinal plant species, has been used extensively as genuine medicinal materials. Certain Epimedium species are endangered due to commercial overexploition, while sustainable application studies, conservation genetics, systematics, and marker-assisted selection (MAS) of Epimedium is less-studied due to the lack of molecular markers. Here, we report a set of expressed sequence tags (ESTs) and simple sequence repeats (SSRs) identified in these ESTs for E. sagittatum.ResultscDNAs of E. sagittatum are sequenced using 454 GS-FLX pyrosequencing technology. The raw reads are cleaned and assembled into a total of 76,459 consensus sequences comprising of 17,231 contigs and 59,228 singlets. About 38.5% (29,466) of the consensus sequences significantly match to the non-redundant protein database (E-value < 1e-10), 22,295 of which are further annotated using Gene Ontology (GO) terms. A total of 2,810 EST-SSRs is identified from the Epimedium EST dataset. Trinucleotide SSR is the dominant repeat type (55.2%) followed by dinucleotide (30.4%), tetranuleotide (7.3%), hexanucleotide (4.9%), and pentanucleotide (2.2%) SSR. The dominant repeat motif is AAG/CTT (23.6%) followed by AG/CT (19.3%), ACC/GGT (11.1%), AT/AT (7.5%), and AAC/GTT (5.9%). Thirty-two SSR-ESTs are randomly selected and primer pairs are synthesized for testing the transferability across 52 Epimedium species. Eighteen primer pairs (85.7%) could be successfully transferred to Epimedium species and sixteen of those show high genetic diversity with 0.35 of observed heterozygosity (Ho) and 0.65 of expected heterozygosity (He) and high number of alleles per locus (11.9).ConclusionA large EST dataset with a total of 76,459 consensus sequences is generated, aiming to provide sequence information for deciphering secondary metabolism, especially for flavonoid pathway in Epimedium. A total of 2,810 EST-SSRs is identified from EST dataset and ~1580 EST-SSR markers are transferable. E. sagittatum EST-SSR transferability to the major Epimedium germplasm is up to 85.7%. Therefore, this EST dataset and EST-SSRs will be a powerful resource for further studies such as taxonomy, molecular breeding, genetics, genomics, and secondary metabolism in Epimedium species.

Development and characterization of genic-SSR markers from different Asia lotus (Nelumbo nucifera) types by RNA-seq

“…pigeonpea (Wang et al, 2010;Dutta et al, 2011). Interestingly, the CCG/CGG and CG/CG motifs have the lowest dominant repeat type in N. nucifera, which is consistent with the results from dicotyledonous plants (Kumpatla and Mukhopadhyay 2005), such as Epimedium sagittatum (Jiang et al, 2012) and radish (Zeng et al, 2010;Jiang et al, 2012). These results showed that the CCG/ CGG motif is the rarest motif in a large number of dicotyledonous plants.…”

“…About 9% (9523 contigs) of the transcriptomic sequences possessed SSR loci. This rate was higher than for Epimedium sagittatum (3.67%), which were identified using the same search parameters (Zeng et al, 2010). The distribution density in N. nucifera is one microsatellite loci per 6.84 kb.…”

“…This might be due to the high GC content and consequent codon usage bias in monocots (Morgante et al, 2002;La Rota et al, 2005). Moreover, the sequences containing CCG/CGG repeats might form potential higher structure, such as hairpins, and thus influence the efficiency and accuracy of RNA splicing (Coleman and Roesser, 1998;Zeng et al, 2010).…”

Transcriptome analysis of potential simple sequence repeat markers in Ammopiptanthus mongolicus

“…Therefore, EST-SSRs have a relatively high transferability compared with genomic SSRs (Wei et al, 2011b). With the development of next-generation sequencing technologies, many EST-SSRs have been found and evaluated in sweet potato (Wang et al, 2010), chickpea (Garg et al, 2011), Epimedium sagittatum (Zeng et al, 2010), Siberian wildrye (Zhou et al, 2016), and multiple other species. However, the results of these studies have shown that SSRs vary in different plant species.…”

Identification of novel and useful EST-SSR markers from de novo transcriptome sequence of wheat (Triticum aestivum L.)

“…These SSR markers can be isolated from both conserved coding regions and non-coding nucleotide sequences of all higher organisms (Sraphet et al, 2011). The development of genomic SSRs, however, is relatively timeconsuming and expensive; but SSRs can alternatively be found in public sequence databases of expressed sequence tags (ESTs) or cDNA (Zeng et al, 2010;Huang et al, 2011;Sraphet et al, 2011;Koelling et al, 2012). Such SSRs are referred to as EST-SSRs.…”