High-throughput analysis and characterization of Astragalus membranaceus transcriptome using 454 GS FLX.

Xiu-Bo Liu,Ling Ma,Ai-Hua Zhang,Lei-Ming Zhang,Xiang-Jun Kong,Wei Ma,Wei-Chao Ren,Jing Jiang,Yan-He Zhang,Jeremy J.W Chen

doi:10.1371/journal.pone.0095831

Abstract

Astragalus membranaceus (Fisch.) Bge (AR), one of the most important medicinal plants in Asia, was found to exhibit various bioactivities. Due to limited genomic and transcriptomic data, the biosynthetic pathway of the major bioactive compound in AR, is currently unclear. In this study, 454 GS FLX technology was employed to produce a substantial expressed sequence tag (EST) dataset from the AR. In all, 742721 high-quality reads from the AR were produced using Roche GS FLX Titanium. A total of 9893 unique sequences were obtained and annotated by a similarity search against the public databases, and involved in the secondary metabolic pathway, which would facilitate deciphering the molecular mechanism of secondary metabolism in AR. The assembled sequences were annotated with gene names and Gene Ontology (GO) terms. GO revealed the unique sequences that could be assigned to 34 vocabularies. In the KEGG mapping, unique sequences were established as associated with 46 biochemical pathways. These results provided the largest EST collections in AR and will contribute to biosynthetic and biochemical studies that lead to drug improvement. With respect to the genes related to metabolism and biosynthesis pathway were also found. Our work demonstrated the utility of 454 GS FLX as a method for the rapid and cost-effective identification of AR transcriptome, and this EST dataset will be a powerful resource for further studies such as taxonomy, molecular breeding, and secondary metabolism in AR.

Highlights

Identification of candidate genes involved in the biosynthetic pathway will significantly contribute to the understanding of the biosynthetic and medicinal chemistry of the plant [1]
Only a small number of Astragalus membranaceus (Fisch.) Bge (AR) expressed sequence tags (EST) have been deposited in GenBank, alignment analysis showed that the majority of ESTs can be found in our 454 sequence collection
To explore the transcriptome of AR and identify genes involved in the metabolism pathway, the 454 GS FLX technology was used in this study to generate substantial transcriptome sequence data

Summary

Introduction

Identification of candidate genes involved in the biosynthetic pathway will significantly contribute to the understanding of the biosynthetic and medicinal chemistry of the plant [1]. Transcriptome sequencing would provide a foundation for detailed studies of gene expression and genetic connectivity with respect to plant secondary metabolism. Various techniques such as microarray, serial analysis of gene expression and massively parallel signature sequencing emerged for high throughput gene expression profiling in the past [5,6]. These techniques are time consuming and become expensive, for the analysis at global level. The 454 GS FLX sequencing technology has made EST-based resources more readily accessible for plant transcriptomes [8]

Methods

Results

Conclusion