Xingyi Wei

Common vetch (Vicia sativa subsp. sativa) is one of the most economically important forage legumes with rich nutritional value and multiple uses. Though the large-scale development and application of microsatellite markers have been conducted in common vetch germplasm evaluation, the investigation and exploitation of intron-length polymorphic (ILP) markers have not been systematically analyzed. In this study, the intron positions of common vetch genome were located by aligning the RNA-Seq sequences of common vetch with Medicago truncatula, soybean (Glycine max), and Arabidopsis thaliana genomic sequences, and used for VsILP marker development. A total of 10,400 markers were generated from 44,582 common vetch unigenes. Out of 300 randomly selected VsILP markers, 283 resulted in successful amplification in common vetch. Among these markers, 40 produced length variation in 30 common vetch accessions, collectively yielding 166 alleles with an average of 4.0 alleles per locus. The polymorphic information content (PIC) values extended from 0.06 to 0.81 with a mean of 0.49. Of the 283 VsILP markers, 84.8% exhibited transferability to leguminous and non-leguminous species. We presented here the first large-scale development of ILP markers in common vetch and their utility in germplasm evaluation and transferability, which will be valuable for further comparative genomic studies, genetic relationship assessments, and marker-assisted breeding of leguminous and non-leguminous species.

Microsatellite (simple sequence repeats, SSRs) marker is one of the most widely used markers in marker-assisted breeding. As one type of functional markers, MicroRNA-based SSR (miRNA-SSR) markers have been exploited mainly in animals, but the development and characterization of miRNA-SSR markers in plants are still limited. In the present study, miRNA-SSR markers for Medicago truncatula (M. truncatula) were developed and their cross-species transferability in six leguminous species was evaluated. A total of 169 primer pairs were successfully designed from 130 M. truncatula miRNA genes, the majority of which were mononucleotide repeats (70.41%), followed by dinucleotide repeats (14.20%), compound repeats (11.24%) and trinucleotide repeats (4.14%). Functional classification of SSR-containing miRNA genes showed that all targets could be grouped into three Gene Ontology (GO) categories: 17 in biological process, 11 in molecular function, and 14 in cellular component. The miRNA-SSR markers showed high transferability in other six leguminous species, ranged from 74.56% to 90.53%. Furthermore, 25 Mt-miRNA-SSR markers were used to evaluate polymorphisms in 20 alfalfa accessions, and the polymorphism information content (PIC) values ranged from 0.39 to 0.89 with an average of 0.71, the allele number per marker varied from 3 to 18 with an average of 7.88, indicating a high level of informativeness. The present study is the first time developed and characterized of M. truncatula miRNA-SSRs and demonstrated their utility in transferability, these novel markers will be valuable for genetic diversity analysis, marker-assisted selection and genotyping in leguminous species.