Noninvasive determination of genome size and ploidy level in fishes by flow cytometry: detection of triploid Poecilia formosa
✍ Scribed by D.K. Lamatsch; C. Steinlein; M. Schmid; M. Schartl
- Publisher
- John Wiley and Sons
- Year
- 2000
- Tongue
- English
- Weight
- 202 KB
- Volume
- 39
- Category
- Article
- ISSN
- 0196-4763
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✦ Synopsis
Background:
In order to understand the evolutionary significance of single triploids among the mostly diploid Poecilia formosa we have developed a simple, noninvasive technique for DNA content and ploidy determination. Methods: From dorsal fin clips of 14 different fish species single cell suspensions were obtained by chopping the material in 2.1% citric acid/0.5% Tween20, passing it through a 0.6-gauge needle and incubating it for 20 min at room temperature (RT) with gentle agitation. After overnight fixation in 70% ethanol, the cells were treated with 1ml 0.5% pepsin/0.1 M HCl for 15 min at RT before adding DAPI to a final volume of 2 ml. The cells were stained for 1-3 h and then analyzed by flow cytometry. Results: We obtained good measurements with CVs ranging from 1.23% to 3.36%. The poeciliid species measured contain from 1.6 to 2.0 pg/nucleus, Oryzias latipes (Medaka) exhibits a nuclear DNA content of 2.2 pg, Danio rerio (zebrafish) 4.6 pg, Tetraodon fluviatilis (freshwater fugu) 0.70 pg. All values except zebrafish are in good agreement with the literature. Conclusions: The identification of living specimens of different ploidy for breeding experiments, behavioral studies and tissue transplantations is now made possible. With slight modifications the method can be extended to a field technique, providing therefore a useful tool for a variety of researchers. Cytometry 39:91-95, 2000.