De novo assembly and transcriptome characterization of an endemic species of Vietnam, Panax vietnamensis Ha et Grushv., including the development of EST-SSR markers for population genetics

Abstract

Background: Understanding the genetic diversity in threatened species that occur in forest remnants is necessary to establish efficient strategies for the species conservation, restoration and management. Panax vietnamensis Ha et Grushv. is medicinally important, endemic and endangered species of Vietnam. However, genetic diversity and structure of population is unknown due to lack of efficient molecular markers. Results: In this study, we employed Illumina HiSeq TM 4000 sequencing to analyze the transcriptomes of P. vietnamensis (roots, leaves and stems). A total of 23,741,783 raw reads were obtained and assembled, from which, 89,271 unigenes with an average length of 598.3191 nt were generated. During functional annotation, 31,686 unigenes were annotated in Gene Ontology categories, Kyoto Encyclopedia of Genes and Genomes pathways, Swiss-Prot database, and Nucleotide Collection (NR/NT) database. In addition, 11,343 expressed sequence tag-simple sequence repeat (EST-SSRs) were detected. From 7,774 primer pairs, 101 were selected for polymorphism validation, in which, 20 primer pairs were successfully amplified to DNA fragments and significant amounts of polymorphism was observed within population. The nine polymorphic microsatellite loci were used to analyze genetic diversity and structure of the natural populations. The obtained results revealed that the shows high levels of genetic diversity in populations, the average observed and expected heterozygosity were H O = 0.422 and H E = 0.479. During the Bottleneck analysis using TPM and SMM models (p < 0.01) shows that targeted population is significantly heterozygote deficient. This suggests sign of bottleneck in all populations. Genetic differentiation among populations was moderate (F ST = 0.133) and indicating limited gene flow (Nm = 1.63). Analysis of molecular variance (AMOVA) showed 63.17% of variation within individuals and 12.45% among populations. These results showed a moderate genetic structure of P. vietnamensis. STRUCTURE analysis and the unweighted pair-group method with arithmetic means (UPGMA) tree revealed strong genetic structure and two genetic clusters related to geographical distances, as well. Conclusion: Our study will assist conservators in future conservation management, breeding, production and habitats restoration of the species. Keywords: Conservation, EST-SSRs; Transcriptome; Panax vietnamensis ; Population genetics