De novo characterization of the Rehmannia glutinosa leaf transcriptome and analysis of gene expression associated with replanting disease

Abstract

The perennial herbaceous plant, Rehmannia glutinosa Libosch, is a traditional Chinese medicine because of the active extracts from its dried tuberous roots. However, R. glutinosa productivity and quality has been seriously affected by replanting (continuous monoculture) disease, which cannot at present be effectively prevented or controlled. Since very little is known about the molecular mechanism of replanting disease, we aimed to investigate transcriptional changes in replanted R. glutinosa leaves and identify genes responding to the disease. Here, we constructed a cDNA library from total RNA isolated from the mixture of leaves of the first year planted (L1) and the second year replanted R. glutinosa (L2) at the tuberous root expansion stage. We generated about 37 million high-quality reads from the cDNA library using deep sequencing and obtained 94,544 distinct sequences by de novo assembly and gap-filling. From this set, a total of 54,490 transcripts containing a complete or partial encoding region was annotated in public protein databases. Based on this resource, we screened differentially expressed genes in the L1 and L2 libraries by the digital gene expression (DGE) technique. Finally, a set of 1,954 genes was found to be differentially expressed in L2. Using bioinformatics and qRT-PCR, the 117 most strongly differentially expressed genes were considered to be prime candidates responsible for replanting disease. Functional analysis of the candidates showed that ethylene signaling was exaggerated and the genes in key metabolism pathways were abnormally expressed in L2. The study provides an important resource for further investigating the cause of replanting disease and developing methods to control or reduce its harmful effects.