Development and characterization of EST-SSR markers in pecan (Carya illinoinensis)

Key message

This study developed and validated a set of polymorphic and cross-transferable EST-SSR markers in pecan. 31 polymorphic primers were qualified for genetic diversity evaluation.

Abstract

Pecan is a famous nut tree with high nutritional value. Nowadays, few EST-SSR markers have been developed for this species. Here, 31,074 protein-coding genes of pecan with 46.43 Mb were employed for EST-SSR identification. In total, 9522 EST-SSR loci were detected from 5752 genes with a distribution frequency of 16.69% of total genes and an average density of one SSR per 4.88 kb. Among them, the di-nucleotide (44.24%) was the most abundant motif, followed by mono-, tri-, tetra-, hexa-, and penta-nucleotide repeats. In addition, a total of 7764 EST-SSR loci were distributed in UTR regions, while the rest of the 1758 were located at CDS regions. Based on the flanked sequences surrounding SSR loci, 3586 primers were successfully developed and 250 pairs were randomly selected for verification using six pecan cultivars. After amplification, 171 (68.40%) primer pairs yielded the expected bands, of which, 159 showed polymorphisms. Among the polymorphic primers, 76 pairs could be cross-amplified in four other Carya species (Carya. laciniosa, C. ovata, C. hunanensis, and C. cathayensis). To further characterize the usefulness of EST-SSRs, 31 polymorphic primers were subjected to assess the genetic diversity and relationship of 40 pecan lines, in which the mean number of alleles per locus and polymorphism information content were 6.90 and 0.70, respectively, suggesting the capacity of the markers to monitor high level of informativeness. Cluster analysis indicated that some accessions in the corresponding subgroup were in good agreement with their genetic backgrounds and morphological traits. The EST-SSR resources offered here constitute a valuable tool for genetic diversity analysis in pecan.