Transcriptome analysis of buds and leaves using 454 pyrosequencing to discover genes associated with the biosynthesis of active ingredients in Lonicera japonica Thunb.

Liu He,Shilin Chen,Xiaolan Xu,Zhiying Sun,Yingjie Zhu,Ruiyang Cheng,Yu’An Bi,Chunfang Li,Jingyuan Song,Haixia Yan,Juan Shen,Ying Li,Zhenzhong Wang,Wei Xiao,Chao Sun,Turgay Unver

doi:10.1371/journal.pone.0062922

Abstract

Background Lonicera japonica Thunb. is a plant used in traditional Chinese medicine known for its anti-inflammatory, anti-oxidative, anti-carcinogenic, and antiviral pharmacological properties. The major active secondary metabolites of this plant are chlorogenic acid (CGA) and luteoloside. While the biosynthetic pathways of these metabolites are relatively well known, the genetic information available for this species, especially the biosynthetic pathways of its active ingredients, is limited.Methodology/Principal FindingsWe obtained one million reads (average length of 400 bp) in a whole sequence run using a Roche/454 GS FLX titanium platform. Altogether, 85.69% of the unigenes covering the entire life cycle of the plant were annotated and 325 unigenes were assigned to secondary metabolic pathways. Moreover, 2039 unigenes were predicted as transcription factors. Nearly all of the possible enzymes involved in the biosynthesis of CGA and luteoloside were discovered in L. japonica. Three hydroxycinnamoyl transferase genes, including two hydroxycinnamoyl-CoA quinate hydroxycinnamoyl transferase genes and one hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferase (HCT) gene featuring high similarity to known genes from other species, were cloned. The HCT gene was discovered for the first time in L. japonica. In addition, 188 candidate cytochrome P450 unigenes and 245 glycosyltransferase unigenes were found in the expressed sequence tag (EST) dataset.ConclusionThis study provides a high quality EST database for L. japonica by 454 pyrosequencing. Based on the EST annotation, a set of putative genes involved in CGA and luteoloside biosynthetic pathways were discovered. The database serves as an important source of public information on genetic markers, gene expression, genomics, and functional genomics in L. japonica.

Highlights

Lonicera japonica Thunb. is a perennial, evergreen, twining vine
Based on the expressed sequence tag (EST) annotation, a set of putative genes involved in chlorogenic acid (CGA) and luteoloside biosynthetic pathways were discovered
51,500 were annotated and all of the unigenes encoding enzymes involved in the CGA and luteoloside biosynthetic pathways were found

Summary

Introduction

It has double-tongued flowers that open white and fade to yellow. L. japonica is believed to be invasive to the ecology of some American countries because of its strong viability [1] It is widely cultivated in China as well as in other Asian countries, such as Japan and Korea, as a commercially valuable plant. Flos Lonicerae Japonicae (FLJ), the dried bud of the L. japonica, has been used for thousands of years in Chinese medicine for its antipyretic, antidotal, and antiinflammatory properties. It has been recorded in the ‘‘Ben Cao Gang Mu’’ (Compendium of Materia Medica) as early as the 17th century. While the biosynthetic pathways of these metabolites are relatively well known, the genetic information available for this species, especially the biosynthetic pathways of its active ingredients, is limited

Methods

Results

Discussion

Conclusion