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Chinese Journal of Oceanology and Limnology

, Volume 33, Issue 4, pp 974–987 | Cite as

Development of a real-time PCR assay (SYBR Green I) for rapid identification and quantification of scyphomedusae Aurelia sp.1 planulae

  • Jianyan Wang (王建艳)
  • Yu Zhen (甄毓)
  • Tiezhu Mi (米铁柱)
  • Zhigang Yu (于志刚)
  • Guoshan Wang (王国善)
Biology
  • 145 Downloads

Abstract

The complicated life cycle of Aurelia spp., comprising benthic asexually-reproducing polyps and sexually-reproducing medusae, makes it hard for researchers to identify and track them, especially for early stage individuals, such as planulae. To solve this problem, we developed a real-time PCR assay (SYBR Green I) to identify planulae in both cultured and natural seawater samples. Species-specific primers targeting Aurelia sp.1 mitochondrial 16S rDNA (mt 16S rDNA) regions were designed. Using a calibration curve constructed with plasmids containing the Aurelia sp.1 mt 16S rDNA fragment and a standard curve for planulae, the absolute number of mt 16S rDNA copies per planula was determined and from that the total number of planulae per sample was estimated. For the field samples, a 100-fold dilution of the sample DNA combined with a final concentration of 0.2 μg/μL BSA in the PCR reaction mixture was used to remove real-time PCR inhibitors. Samples collected in Jiaozhou Bay from July to September 2012 were subsequently analyzed using this assay. Peak Aurelia sp.1 planula abundance occurred in July 2012 at stations near Hongdao Island and Qingdao offshore; abundances were very low in August and September. The real-time PCR assay (SYBR Green I) developed here negates the need for traditional microscopic identification, which is laborious and time-consuming, and can detect and quantify jellyfish planulae in field plankton samples rapidly and specifically.

Keywords

Aurelia sp.1 16S rDNA planulae real-time PCR jellyfish blooms 

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Copyright information

© Chinese Society for Oceanology and Limnology, Science Press and Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Jianyan Wang (王建艳)
    • 1
  • Yu Zhen (甄毓)
    • 2
    • 3
  • Tiezhu Mi (米铁柱)
    • 2
    • 3
  • Zhigang Yu (于志刚)
    • 4
    • 5
  • Guoshan Wang (王国善)
    • 1
  1. 1.College of Marine Life SciencesOcean University of ChinaQingdaoChina
  2. 2.College of Environmental Science and EngineeringOcean University of ChinaQingdaoChina
  3. 3.Key Laboratory of Marine Environment and Ecology, Ministry of EducationOcean University of ChinaQingdaoChina
  4. 4.College of Chemistry and Chemical EngineeringOcean University of ChinaQingdaoChina
  5. 5.Key Laboratory of Marine Chemistry Theory and Technology, Ministry of EducationOcean University of ChinaQingdaoChina

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