Flow cytometric characterization of the chlorophyll contents and size distributions of plant protoplasts.

David W Galbraith, K. R. Harkins, R. A. Jefferson

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

We have employed flow cytometry for the characterization of populations of protoplasts prepared from tobacco (Nicotiana tabacum) leaf tissues. We first investigated the possibility of using flow cytometric analysis of the emission of chlorophyll autofluorescence for measurement of the chlorophyll contents of leaf protoplasts. Defined numbers of leaf protoplasts were sorted according to different, nonoverlapping windows placed on the one-dimensional histograms of chlorophyll autofluorescence emission. The amounts of cellular chlorophyll were measured in cell-free extracts of these sorted protoplasts using fluorometry. A high degree of correlation (r2 = 0.983) was observed between these two parameters. We then examined the distribution of protoplast diameters in these protoplast populations through the use of pulse-width time-of-flight (TOF) analysis. Through sorting of protoplasts using a series of narrow, nonoverlapping TOF windows, we were able to demonstrate that the TOF parameter was linearly correlated with protoplast diameter, over the range of 15-55 micron (r2 greater than 0.99). We also compared the use of fluorescein diacetate (FDA) fluorochromasia and chlorophyll autofluorescence as the source of fluorescent signals for TOF analysis. We found that the presence of chloroplasts introduced distortions into the measurement of apparent size afforded by TOF analysis of FDA fluorochromasia. These results are discussed in terms of the application of techniques of flow analysis and sorting for the measurement of gene expression within the various different cell types found in plant tissues and organs.

Original languageEnglish (US)
Pages (from-to)75-83
Number of pages9
JournalCytometry
Volume9
Issue number1
StatePublished - Jan 1988
Externally publishedYes

Fingerprint

Plant Dispersal
Protoplasts
Chlorophyll
Tobacco
Fluorometry
Chloroplasts
Cell Extracts
Population
Flow Cytometry
Gene Expression

ASJC Scopus subject areas

  • Biophysics
  • Cell Biology
  • Endocrinology
  • Hematology
  • Pathology and Forensic Medicine

Cite this

Flow cytometric characterization of the chlorophyll contents and size distributions of plant protoplasts. / Galbraith, David W; Harkins, K. R.; Jefferson, R. A.

In: Cytometry, Vol. 9, No. 1, 01.1988, p. 75-83.

Research output: Contribution to journalArticle

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