Abstract
BackgroundLychnis kiusiana Makino is an endangered perennial herb native to wetland areas in Korea and Japan. Despite its conservational and evolutionary significance, population genetic resources are lacking for this species. Next-generation sequencing has been accepted as a rapid and cost-effective solution for the identification of microsatellite markers in nonmodel plants.ResultsUsing Illumina HiSeq 2000 sequencing technology, we assembled 67,498,600 reads into 91,900 contigs and identified 11,403 microsatellite repeat motifs in 9563 contigs. A total of 4510 microsatellite-containing transcripts had Gene Ontology (GO) annotations, and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis identified 124 pathways with significant scores. Many microsatellites in the L. kiusiana leaf transcriptome were linked to genes involved in the plant response to light intensity, salt stress, temperature stimulus, and nutrient and water deprivation. A total of 12,486 single-nucleotide polymorphisms (SNPs) were identified on transcripts harboring microsatellites. The analysis of nucleotide substitution rates for 2389 unigenes indicated that 39 genes were under strong positive selection. The primers of 6911 microsatellites were designed, and 40 of 50 selected primer pairs were consistently and successfully amplified from 51 individuals. Twenty-five of these were polymorphic, and the average number of alleles per SSR locus was 6.96, with a range from 2 to 15. The observed and expected heterozygosities ranged from 0.137 to 0.902 and 0.131 to 0.827, respectively, and locus-specific FIS estimates ranged from − 0.116 to 0.290. Eleven of the 25 primer pairs were successfully amplified in three additional species of Lychnis: 56% in L. wilfordii, 64% in L. cognata and 80% in L. fulgens.ConclusionsThe transcriptomic SSR markers of Lychnis kiusiana provide a valuable resource for understanding the population genetics, evolutionary history, and effective conservation management of this species. Furthermore, the identified microsatellite loci linked to the annotated genes should be useful for developing functional markers of L. kiusiana. The developed markers represent a potentially valuable source of transcriptomic SSR markers for population genetic analyses with moderate levels of cross-taxon portability.
Highlights
Lychnis kiusiana Makino is an endangered perennial herb native to wetland areas in Korea and Japan
Transcriptomic sequencing has been applied for mining microsatellite markers in related species, i.e., Silene vulgaris [22], and the results indicated that transcriptomic Simple sequence repeat (SSR) provide potential for genomic and population genetic approaches
Ten tri-nucleotide motif types were identified with AAT/ATT accounting for approximately 21.4%, while CCG/CGG only accounted for 1.7% (Additional file 2: Figure S1)
Summary
Lychnis kiusiana Makino is an endangered perennial herb native to wetland areas in Korea and Japan. Next-generation sequencing has been accepted as a rapid and cost-effective solution for the identification of microsatellite markers in nonmodel plants. Microsatellites, or simple sequence repeats (SSRs), are tandem repeats of mono- to hexa-nucleotide sequence motifs that generally vary in length between five and 40 repeats. They are codominant, abundant, multiallelic and locus-specific markers and are found at high frequency in eukaryotic genomes [1, 2]. Next-generation sequencing (NGS) has been accepted as a rapid and cost-effective solution for the identification of microsatellite markers in nonmodel plants [5, 6]. Transcriptomic SSRs may play an important role in adaptive evolution by generating genetic variation [9]
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