Development of SSR markers and genetic diversity analysis in enset (Ensete ventricosum (Welw.) Cheesman), an orphan food security crop from Southern Ethiopia

Temesgen Magule Olango, Bizuayehu Tesfaye, Mario Augusto Pagnotta, Mario Enrico Pè, Marcello Catellani

Enset (Ensete ventricosum (Welw.) Cheesman; Musaceae) is a multipurpose drought-tolerant food security crop with high conservation and improvement concern in Ethiopia, where it supplements the human calorie requirements of around 20 million people. The crop also has an enormous potential in other regions of Sub-Saharan Africa, where it is known only as a wild plant. Despite its potential, genetic and genomic studies supporting breeding programs and conservation efforts are very limited. Molecular methods would substantially improve current conventional approaches. Here we report the development of the first set of SSR markers from enset, their cross-transferability to Musa spp., and their application in genetic diversity, relationship and structure assessments in wild and cultivated enset germplasm. SSR markers specific to E. ventricosum were developed through pyrosequencing of an enriched genomic library. Primer pairs were designed for 217 microsatellites with a repeat size > 20 bp from 900 candidates. Primers were validated in parallel by in silico and in vitro PCR approaches. A total of 67 primer pairs successfully amplified specific loci and 59 showed polymorphism. A subset of 34 polymorphic SSR markers were used to study 70 both wild and cultivated enset accessions. A large number of alleles were detected along with a moderate to high level of genetic diversity. AMOVA revealed that intra-population allelic variations contributed more to genetic diversity than inter-population variations. UPGMA based phylogenetic analysis and Discriminant Analysis of Principal Components show that wild enset is clearly separated from cultivated enset and is more closely related to the out-group Musa spp. No cluster pattern associated with the geographical regions, where this crop is grown, was observed for enset landraces. Our results reaffirm the long tradition of extensive seed-sucker exchange between enset cultivating communities in Southern Ethiopia. The first set of genomic SSR markers were developed in enset. A large proportion of these markers were polymorphic and some were also transferable to related species of the genus Musa. This study demonstrated the usefulness of the markers in assessing genetic diversity and structure in enset germplasm, and provides potentially useful information for developing conservation and breeding strategies in enset.