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Planta

, Volume 249, Issue 4, pp 1229–1237 | Cite as

Transcriptome analysis of 1- and 3-year-old Panax notoginseng roots and functional characterization of saponin biosynthetic genes DS and CYP716A47-like

  • Jian Li
  • Lan Ma
  • Shuting Zhang
  • Cailian Zuo
  • Na Song
  • Shusheng Zhu
  • Jinsong WuEmail author
Original Article

Abstract

Main conclusion

Transcriptome analysis revealed high expression of saponin biosynthetic genes may account for highly accumulated saponins in 3-year-old Panax notoginseng roots and DS and CYP716A47 - like were functionally verified by transgenic tobacco.

Panax notoginseng is a well-known traditional medical herb that contains bioactive compounds known as saponins. Three major dammarene-type triterpene saponins including R1, Rb1, and Rg1 were found to be highly accumulated in the roots of 3-year-old plants when compared to those of 1-year-old plants. However, the underlying cellular mechanism is poorly understood. In this study, transcriptome analysis revealed that most genes involved in saponin biosynthesis in P. notoginseng roots augmented during their growth periods. The analysis of the KEGG pathway indicated that the primary metabolism, cell growth, and differentiation were less active in the roots of 3-year-old plant; however, secondary metabolisms were enhanced, thus providing molecular evidence for the harvesting of P. notoginseng roots in the 3rd year of growth. Furthermore, the functional role of DS and CYP716A47-like, two of the candidate genes involved in saponin biosynthesis isolated from P. notoginseng, were verified via overexpression in cultivated tobacco. Approximately, 0.325 µg g−1 of dammarenediol-II and 0.320 µg g−1 of protopanaxadiol were recorded in the dry leaves of transgenic tobacco overexpressed with DS and both DS and CYP716A47-like, respectively. This study provides insights into the molecular mechanisms for saponin accumulation in P. notoginseng roots during its growth period and paves a promising way to produce dammarenediol-II and protopanaxadiol via transgenic techniques.

Keywords

1- and 3-year-old Panax notoginseng Dammarene-type triterpene saponins RNA-seq Transgenic tobacco 

Abbreviations

HPLC

High-performance liquid chromatography

MVA

Mevalonic acid

AACT

Acetyl-CoA acetyltransferase

HMGS

3-Hydroxy-3-methylglutaryl coenzyme-A synthase

HMGR

HMG-CoA reductase

MVK

Mevalonate kinase

PMK

Phosphomevalonate kinase

MVD

Mevalonate diphosphate decarboxylase

IDI

Isopentenyl diphosphate isomerase

GGPS

Geranylgeranyl pyrophosphate synthase

FPS

Farnesyl diphosphate synthase

SS

Squalene synthase

SE

Squalene epoxidase

DS

Dammarenediol-II synthase

β-AS

Beta-amyrin synthase

CS

Cycloartenol synthase

CDS

Coding sequence

DEG

Differentially expressed genes

TFs

Transcription factors

WT

Wild type

Notes

Acknowledgements

We thank Biological Technology Open Platform of Kunming Institute of Botany, the Chinese Academy of Sciences for greenhouse services. This project is supported by the Major Science and Technique Programs in Yunnan Province (No. 2016ZF001), and 100-Oversea-Top-Talents Recruitment plan of Yunnan.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

425_2018_3083_MOESM1_ESM.pdf (112 kb)
Supplementary material 1 (PDF 112 kb)
425_2018_3083_MOESM2_ESM.xlsx (24 kb)
Supplementary material 2 (XLSX 23 kb)
425_2018_3083_MOESM3_ESM.tif (130 kb)
Supplementary material 3 Detection of introduced genes in transgenic tobacco overexpressing DS and CYP716A47-like. a Confirmation of the introduced genes DS by real-time PCR. b Expression of DS and CYP716A47-like in leaves of wild-type Nicotiana tobacum (WT) and DS/CYP transgenic lines (the DS and CYP716A47-like co-overexpressed lines) were detected by real-time PCR. Data are mean values with the standard deviation obtained from three independent plants. Not Detected (ND) means under detection limits (TIFF 129 kb)
425_2018_3083_MOESM4_ESM.tif (41 kb)
Supplementary material 4 Relative transcripts of genes GUS and DS in leaves of wild-type Nicotiana tobacum (WT), empty vector transgenic line (EV) and DS transgenic line 1 (DS 1#) were detected by real-time PCR. Data are mean values with the standard deviation obtained from three independent plants. The asterisks indicate significant differences between WT and other samples (Paired t test: **P < 0.01). Not Detected (ND) means under detection limits (TIFF 41 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Yunnan Key Laboratory for Wild Plant ResourcesKunming Institute of Botany, Chinese Academy of SciencesKunmingChina
  2. 2.University of Chinese Academy of SciencesBeijingChina
  3. 3.Key Laboratory of Agro-Biodiversity and Pest Management of Education Ministry of China, State Key Laboratory for Conservation and Utilization of Bio-Resources in YunnanYunnan Agricultural UniversityKunmingChina

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