Construction of an SNP linkage map using genotyping-by-sequencing in Capsicum chinense

Abstract

Construction of linkage map is important for genetic studies and marker-assisted breeding programs. Recent advances in next-generation sequencing technologies allow the rapid and efficient generation of high-density linkage maps. In this study, we used a modified genotyping-by-sequencing (GBS) protocol involving two restriction enzymes (PstI-MseI). A total of 87 F2 individuals derived from a cross of C. chinense Habanero and C. chinense Jolokia were used to construct an SNP linkage map. To imputate missing data, a novel method combining sliding window and bin mapping was used to obtain reliable genotype data. A total of 1,000 bin markers selected from 217,152 SNPs were used for construction of a linkage map. The total genetic distance was about 3654.1 cM, and the average distance between bin markers was 3.9 cM. These bin markers were evenly distributed over 14 linkage groups, corresponding to 12 pepper chromosomes. This research shows that GBS together with the sliding window imputation method enabled us to rapidly develop a genetic map in an F2 generation. Using the linkage map, QTL mapping was performed for capsaicinoids content in the placenta and pericarp tissues. Also, we analyzed Pun1 expression to test whether capsaicinoids are synthesized in the pericarp. The results showed that only Jolokia showed Pun1 expression whereas Habanero and F1 hybrid showed no expression in pericarp. Distribution of capsaicinoids content in the pericarp of F2 plants also indicated that capsaicinoids might be regulated by a recessive gene(s). When QTL analysis was performed using the GBS linkage map and capsaicinoids content data, one QTL was identified for capsaicinoids in the placenta. Six QTLs were detected on chromosome 6 and 9 for capsaicinoids content in the pericarp. Bin markers linked to the two major QTLs will be used for selection plants containing high capsaicinoids content.