Development of SNP markers using RNA-seq technology and tetra-primer ARMS-PCR in sweetpotato

Abstract

The information of single nucleotide polymorphisms (SNPs) is quite unknown in sweetpotato. In this study, two sweetpotato varieties (Xushu 18 and Xu 781) were sequenced by Illumina technology, as well as de novo transcriptome assembly, functional annotation, and in silico discovery of potential SNP molecular markers. Tetra-primer Amplification Refractory Mutation System PCR (ARMS-PCR) is a simple and sufficient method for detecting different alleles in SNP locus. Total 153 sets of ARMS-PCR primers were designed to validate the putative SNPs from sequences. PCR products from 103 sets of primers were different between Xu 781 and Xushu 18 via agarose gel electrophoresis, and the detection rate was 67.32%. We obtained the expected results from 32 sets of primers between the two genotypes. Furthermore, we ascertained the optimal annealing temperature of 32 sets of primers. These SNPs might be used in genotyping, QTL mapping, or marker-assisted trait selection further in sweetpotato. To our knowledge, this work was the first study to develop SNP markers in sweetpotato by using tetra-primer ARMS-PCR technique. This method was a simple, rapid, and useful technique to develop SNP markers, and will provide a potential and preliminary application in discriminating cultivars in sweetpotato.

Keywords

• sweetpotato
• SNPs
• RNA-seq
• tetra-primer ARMS-PCR