Development of Polymorphic SSR Markers in Chinese Water Chestnut Based on RAD-seq

Abstract

Chinese water chestnut ( Eleocharis dulcis ) is an important characteristic vegetable that grows in shallow waters. In order to develop simple sequence repeat (SSR) molecular markers for genetics research on Eleocharis dulcis , simplified genome sequencing by restriction-site associated DNA sequencing (RAD-seq), SSR search and primer design were performed. A total of 5039 SSR loci were detected, from which 4 137 pairs of primer were designed. Among them, the proportion of tri-nucleotides motifs was the highest (1 894), accounting for 45.89% of the total loci; followed by di-nucleotide with 1 406, accounting for 34.07%.. A total of 100 pairs of randomly selected primers were verified. 93% of the primers were successfully amplified. To assess the polymorphism of these primers, two varieties of Chinese water chestnuts were used to amplify the SSR marker using 83 pairs of primers. The 83 pairs of primers generated a total of 232 fragments. A total of 128 fragments were polymorphic. 60 pairs of primers out of 83 pairs generated at least one polymorphic band, and the ratio of polymorphic primers was 72.28%. These SSR markers developed from RAD tags were proved to be effective, which will provide a useful tool for genetic diversity research and molecular breeding in Chinese water chestnuts.