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Molecular Biology Reports

, Volume 40, Issue 2, pp 2035–2044 | Cite as

Molecular cloning and characterization of three isoprenyl diphosphate synthase genes from alfalfa

  • Yan Sun
  • Ruicai Long
  • Junmei Kang
  • Tiejun Zhang
  • Ze Zhang
  • He Zhou
  • Qingchuan YangEmail author
Article

Abstract

Isoprenoid is the precursor for the biosynthesis of saponins, abscisic acid, gibberellins, chlorophylls and many other products in plants. Saponins are an important group of bioactive plant natural products. The alfalfa (Medicago sativa L.) saponins are glycosides of different triterpene aglycones and possess many biological activities. We isolated three genes (MsFPPS, MsGPPS and MsGGPPS) encoding isoprenyl diphosphate synthases (IDS) from alfalfa via a homology-based PCR approach. The enzyme activity assay of purified recombined MsFPPS and MsGGPPS expressed in Escherichia coli indicated that they all had IDS activity. Expression analysis of the three genes in different alfalfa tissues using real time PCR displayed that they were expressed in all tissues although they had a different expression patterns. MsFPPS and MsGPS displayed a significant increase in transcript level in response to methyl jasmonate, but the transcript level of MsGGPPS decreased obviously. To elucidate the functions of the three IDSs, their overexpression driven by a constitutive cauliflower mosaic virus-35S promoter in tobacco plants was applied and analyzed. The T0 transgenic plants of MsFPPS showed high levels of squalene content when compared with control. However, no differences were detected in T0 transgenic plants of MsGPPS and MsGGPPS. In addition, the overexpression of MsFPPS induced senescence response in transgenic plant leaves. This result may indicate that MsFPPS performs a role not only in phytosterol and triterpene biosynthesis, but also in growth regulation.

Keywords

Alfalfa Isoprenyl diphosphate synthases Methyl jasmonate Recombinant proteins Enzyme activity Overexpression 

Notes

Acknowledgments

This work was supported by Industry System of Modern Agriculture Construction Funds, China (No. CARS-35).

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Copyright information

© Springer Science+Business Media Dordrecht 2012

Authors and Affiliations

  • Yan Sun
    • 1
  • Ruicai Long
    • 2
  • Junmei Kang
    • 3
  • Tiejun Zhang
    • 3
  • Ze Zhang
    • 2
  • He Zhou
    • 1
  • Qingchuan Yang
    • 3
    Email author
  1. 1.College of Animal Science and TechnologyChina Agriculture UniversityBeijingChina
  2. 2.Institute of Agriculture and Life ScienceChongqing UniversityChongqingChina
  3. 3.Institute of Animal SciencesChinese Academy of Agricultural SciencesBeijingChina

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