Objective SSRs in the transcriptome of Nymphaea Paul Stetson flowers were studied to generate new markers for evaluating germplasms and facilitating breeding of tropical waterlilies.
Method SSR loci were retrieved from the transcriptomes of floral pistils, stamens, and petals of Nymphaea Paul Stetson using MISA. Characteristics of the loci were analyzed by Excel, and primers designed by Primer 3.0 and screened by TP-M13-SSR PCR.
Result There were 12365 SSR loci found in the 39079 unigenes of the transcriptome at the frequency of 31.64% averaging one SSR locus per 5.79 kb. Most of the SSR loci had dinucleotide repeat motifs comprising 71.85% of total with AG/CT being the dominant unit that made up 61.34% of the motifs. Trinucleotide repeat motifs accounted for 26.10% of the sites with AAG/CTT being dominant at 8.30%. The repeating frequency was 5–20 times with a sequence of 12–30 bp averaging 18.38 bp long. Of the 9212 pairs of primers designed, 100 were randomly selected for a validation by PCR amplification to arrive at 9 pairs with high polymorphism being used as the markers. Subsequently, the 12 germplasms were clustered into 3 branches under a genetic similarity coefficient of 0.7375.
Conclusion The SSR loci in the Nymphaea Paul Stetson transcriptome were high on distribution frequency, rich in diversity, greatly polymorphic, and desirable for applications. The 9 pairs of SSR primers identified in this study extended the existing marker repertoire facilitating effective germplasm differentiation on waterlilies.
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