Life Cycle Analysis of Unicellular Algae

Bogdan I. Gerashchenko1, Toshiyuki Takahashi2, Toshikazu Kosaka3, Hiroshi Hosoya3

1 R.E. Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology, Kyiv, Ukraine, 2 Miyakonojo National College of Technology, Miyakonojo, Japan, 3 Hiroshima University, Hiroshima, Japan
Publication Name:  Current Protocols in Cytometry
Unit Number:  Unit 11.19
DOI:  10.1002/0471142956.cy1119s52
Online Posting Date:  April, 2010
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Abstract

Unicellular green alga is a very convenient object for flow cytometric characterization. Flow cytometry has been proposed as a quick and reliable tool for studying life cycle and growth of unicellular algae. Cell size of vegetating algae can be monitored in association with their DNA and endogenous chlorophyll content. Cells of interest (e.g., group of cells of a certain stage of the life cycle) in an asynchronously proliferating cell population can be sorted out for further microscopical or molecular biology studies. This methodological approach can be helpful for researchers who are interested in algal proliferation. Curr. Protoc. Cytom. 52:11.19.1‐11.19.6. © 2010 by John Wiley & Sons, Inc.

Keywords: unicellular algae; Chlorella; light scatter; DNA content; chlorophyll content; life cycle; flow cytometry

     
 
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Table of Contents

  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

Basic Protocol 1:

  Materials
  • Unicellular algae (e.g., Chlorella sp.)
  • Phosphate‐buffered saline (PBS; appendix 2A)
  • Ethanol (≥99.5% purity)
  • DNA staining solution (see recipe)
  • Formaldehyde (methanol free)
  • DAPI
  • Culture cabinet (thermostated at 24°C) with a shaker inside generating 180 reciprocal movements per min and with a light source producing the light of ∼2000 lux
  • 5‐ml centrifuge tubes
  • Hemacytometer
  • Centrifuge for concentration of algal suspensions and for washing steps
  • Refrigerator for incubation at 4°C
  • 37°C incubator
  • 12 × 75–mm polystyrene tubes
  • Flow cytometer/cell sorter equipped with an argon‐ion laser (488 nm)
  • Software for analyzing data obtained by flow cytometry (e.g., CELLQuest software from Becton‐Dickinson Immunocytometry Systems)
  • Microscopes (bright field and fluorescence)
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Figures

Videos

Literature Cited

Literature Cited
   Gerashchenko, B.I., Nishihara, N., Ohara, T., Tosuji, H., Kosaka, T., and Hosoya, H. 2000. Flow cytometry as a strategy to study the endosymbiosis of algae in Paramecium bursaria. Cytometry 41:209‐215.
   Gerashchenko, B.I., Kosaka, T., and Hosoya, H. 2001. Growth kinetics of algal populations exsymbiotic from Paramecium bursaria by flow cytometry measurements. Cytometry 44:257‐263.
   Gerashchenko, B.I., Kosaka, T., and Hosoya, H. 2002. Optical compartmentation of vegetating algae species as a basis for their growth‐specific characterization. Cytometry 48:153‐158.
   Kadono, T., Kawano, T., Hosoya, H., and Kosaka, T. 2004. Flow cytometric studies of the host‐regulated cell cycle in algae symbiotic with green paramecium. Protoplasma 223:133‐141.
   McAuley, P.J. and Muscatine, L. 1986. The cell cycle of symbiotic Chlorella. IV. DNA content of algae slowly increases during host starvation of green hydra. J. Cell Sci. 85:73‐84.
   Nishihara, N., Horiike, S., Takahashi, T., Kosaka, T., Shigenaka, Y., and Hosoya, H. 1998. Cloning and characterization of endosymbiotic algae isolated from Paramecium bursaria. Protoplasma 203:91‐99.
   Takahashi, T., Shirai, Y., Kosaka, T., and Hosoya, H. 2007. Arrest of cytoplasmic streaming induces algal proliferation in green paramecia. PLoS ONE 12:e1352.
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