Development of SSR and gene-targeted markers for construction of a framework linkage map of Catharanthus roseus

Catharanthus roseus is a plant of great medicinal importance, yet inadequate knowledge of its genome structure and the unavailability of genomic resources have been major impediments in the development of improved varieties. The aims of this study were to develop co-dominant sequence-tagged microsatellite sites (STMS) and gene-targeted markers (GTMs) and utilize them for the construction of a framework intraspecific linkage map of C. roseus.

For simple sequence repeat (SSR) isolation, a genomic library enriched for (GA)(n) repeats was constructed from C. roseus 'Nirmal' (CrN1). In addition, GTMs were also designed from 12 genes of the TIA (terpenoid indole alkaloid) pathway - the medicinally most significant pathway in C. roseus. An F(2) mapping population was also generated by crossing two diverse accessions of C. roseus CrN1 (Nirmal)×CrN82 (Kew).

A new set of 314 STMS markers and 64 GTMs were developed in this study. A segregating F(2) mapping population consisting of 111 F(2) individuals was generated. For generating the linkage map, a set of 423 co-dominant markers (378 newly developed and 45 published earlier) were screened for polymorphism between the parental genotypes, of which 134 were identified to be polymorphic. A total of 114 markers were mapped on eight linkage groups that spanned a 632·7 cM region of the genome with an average marker distance of 5·55 cM. Further, the mechanism of hypervariability at the gene-targeted loci was investigated at the sequence level.

For the first time, a large array of STMS markers and GTMs was generated in the model medicinal plant C. roseus. Moreover, the first microsatellite marker-based linkage map was described in this study. Together, these will serve as a foundation for future genomics studies related to quantitative trait loci analysis and molecular breeding in C. roseus.