De novo transcriptome sequencing-based discovery and expression analyses of verbascoside biosynthesis-associated genes in Rehmannia glutinosa tuberous roots

摘要

Verbascoside is a phenylethanoid glycoside found in many plant species, and possessing some pharmacologically beneficial activities for human health. The tentative pathway and early steps of verbascoside biosynthesis have been documented, but key enzymes and their corresponding genes await discovery. We aimed to discover and analyze verbascoside biosynthesis-associated genes. The combined Rehmannia glutinosa tuberous root transcriptome was sequenced de novo. Some verbascoside biosynthesis-associated genes were discovered by the informatics method, validated by RT-PCR and analyzed by qRT-PCR. 149.8 million sequencing reads were generated and assembled de novo, yielding 96,961 final unigenes. Of them, 11,848 unigenes were mapped to 280 KEGGpathways, and 19 genes were assigned to the verbascoside biosynthesis pathway by comparing KEGG pathways and the tentative pathway. Eight genes were validated using RT-PCR in the dataset; qRT-PCR analysis showed that they were expressed in tuberous roots and leaves of both high and low verbascoside R. glutinosa cultivars, and their expressions were greater in the high verbascoside cultivar than in the low one, in old tuberous roots than in young ones, and in leaves than in tuberous roots. The differences in their relative expression levels suggest decisive roles for these enzymes in verbascoside biosynthesis. The results will accelerate the understanding of the genetic basis of verbascoside biosynthesis and genetic engineering breeding in R. glutinosa.