Abstract
As a native in Southeast Asia, Sagittaria trifolia L. var. sinensis (arrowhead) is an aquatic perennial herb of Alismataceae family. For the aim of discovering genes and development molecular markers, high-throughput transcriptome sequencing is applied to generate enormous transcript sequences from arrowhead leaf generate. Through trimmed and assembled, 51, 836 contigs were obtained with mean length of 680 bp. Sequence similarity analyses against four public databases (NR, GO, KEGG, KOG) found 23, 595 contigs that could be annotated with gene descriptions, conserved protein domains, or gene ontology terms. In this study, 3, 861 simple sequence repeat (SSR) markers of arrowhead were identified in 3, 174 contigs of transcriptome data,and 2, 476 primer pairs were designed for marker development. Of these, 100 primers were randomly selected, synthesized and used for validation of the amplification and assessment of polymorphisms in 79 arrowhead accessions. 78 loci reliably amplified a clear single band of expexted size, and 68 loci were scoreable and polymorphic with 2 to 11 alleles per locus. The polymorphism information content valued ranged from 0.01 to 0.84. The SSR markers developed from arrowhead, could be successfully applied to Sagittaria pygmaea, with a transferability rate of 91.14% (72/79). The phylogenetic tree was also constructed to analyse the genetic diversity in arrowhead accessions. The large number of SSR markers developed in this study will be useful in the researches of population genetics, genetic diversity, and germplasm characterization in arrowhead.
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We thank to the editor and reviewer for their constructive suggestions and comments. This research was supported by National Key Technologies R&D Program (No. 2012BAD27B01).
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Electronic Supplementary Material
Supplementary file 1
– Primer sequences for SSR loci. SSR primers were designed using Primer 3.0. The sequence ID, SSR type, SSR motif type, motif size, location of the SSR, location of primers, primer sequences, length of primers, melting temperature, and product size are provided for each locus. (XLS 682 kb)
Supplementary file 2
– Characterization of 100 primer pairs amplification in 79 individual plants. Primer name, sequence ID, repeat motif, sequence of primers, melting temperature and expected size (bp) are indicated. (XLS 45 kb)
Supplementary file 3
– Details of 79 validated EST-SSR markers and predicted functions of their genes, based on BLASTX. Primer name, sequence ID, repeat motif, expected size (bp), objective size (bp), number of varieties from which it was successfully amplified, N. alleles, PIC value, NR_Function, and KOG_Function are indicated. (XLSX 21 kb)
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You, Y., Huang, X., Liu, H. et al. Leaf Transcriptome Analysis and Development of EST-SSR Markers in Arrowhead (Sagittaria trifolia L. Var. Sinensis). Tropical Plant Biol. 13, 189–200 (2020). https://doi.org/10.1007/s12042-019-09242-2
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DOI: https://doi.org/10.1007/s12042-019-09242-2