Novel Microsatellite Markers Derived from Arachis pintoi Transcriptome Sequencing for Cross-Species Transferability and Varietal Identification

Forage peanut ( Arachis pintoi ) is an important leguminous forage that has gained popularity due to increased livestock productivity. Furthermore, the species helps with soil fertility and the restoration of degraded areas. However, A . pintoi has a limited number of molecular markers. The objective of this study was to create and characterize gene-derived microsatellite markers as well as to test their transferability to the peanut ( Arachis hypogaea ) and other six wild Arachis species. A total of 4461 putative simple sequence repeats (SSR) were identified, and PCR primer pairs were designed for 999 SSR regions after filtering out primers with the same annealing site and searching for sequences related to open reading frames (ORFs). The dinucleotide motif was the most common (628; 62.86%). For validation, 186 primer pairs were chosen at random, of which 63 (33.87%) were polymorphic, with an average of 7.37 alleles per locus. Polymorphic information content (PIC = 0.70) and discriminatory power ( D = 0.80) were both high on average. The functional annotation discovered 120 sequences that were assigned to 87 gene ontology functional groups divided into three main categories: molecular function (27 sub-categories), cellular components (21 sub-categories), and biological process (39 sub-categories). Thirty-three SSRs were tested for transferability to peanut and six other wild Arachis species, resulting in variable cross-species amplification (63.64 to 100%). Here, we present the first gene-derived SSR for A . pintoi . These new informative microsatellites may be linked to agronomically important genes to be used in genetic studies.