De novo assembly of transcriptome from Rhododendron latoucheae Franch. using Illumina sequencing and development of new EST-SSR markers for genetic diversity analysis in Rhododendron

Abstract

Rhododendron is universally known for its showy flower, and Rhododendron latoucheae Franch. is an adaptable and wide altitude range species of Rhododendron, with beautiful fragrant flowers and medicinal functions. However, genetic studies of Rhododendron have been hindered by limited genomic resources and genetic markers. In this study, a large dataset composed of 80,660 unigenes derived from the R. latoucheae transcriptome was assembled. Among these unigenes, 34,867 unigenes (43.22%) were annotated against public protein databases. In addition, a total of 14,415 EST-SSRs were identified from 16,019 unigene sequences with an average of one SSR per 2.87 kb. Di-nucleotide SSRs were the most abundant motif (80.03%), followed by tri- (18.82%), tetra- (0.91%), penta- (0.13%), and hexa-nucleotide (0.11%) repeats. Further, 200 primer pairs with more than six tandem repeats were selected, out of which 58 primer pairs amplified successfully and revealed polymorphism among six Rhododendron species. Among these 58 primer pairs, 37 primer pairs were polymorphic in 37 Rhododendron species and 30 R. latoucheae individuals from two national Chinese populations. A total of 382 alleles were identified, with an average of 10.3 alleles per locus. The phylogenetic tree, genetic structure analysis, and principal coordinate analysis demonstrated that major clusters corresponded to known phylogenetic trees and geographic distribution. These transcriptome data will facilitate gene discovery and functional genomic studies in Rhododendron, and the newly developed EST-SSR markers can be applied across multiple Rhododendron species and used for the evaluation of genetic relationships in Rhododendron.