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A simple and sensitive DNA assay for plant extracts
G R Baer1, S P Meyers, W T Molin
1Department of Agronomy, University of Wisconsin, Madison, Wisconsin 53706.
Plant Physiology
|October 1, 1982
Summary
A new fluorimetric method accurately quantifies DNA in plant extracts using the DAPI dye. This method is more sensitive and faster than the diphenylamine assay for DNA determination.
Area of Science:
- Biochemistry
- Molecular Biology
- Plant Science
Background:
- Accurate DNA quantification is crucial for plant molecular biology research.
- Existing methods like the diphenylamine assay have limitations in sensitivity and time efficiency.
Purpose of the Study:
- To develop a sensitive and efficient fluorimetric method for DNA determination in plant extracts.
- To compare the novel fluorimetric method with the traditional diphenylamine assay.
Main Methods:
- Utilized 4',6'-diamidino-2-phenylindole (DAPI) dye for DNA complex formation and fluorescence measurement.
- Optimized DNA recovery and histone dissociation using 2.0 M NaCl.
- Removed interfering pigments and proteins with chloroform treatment to reduce fluorescence quenching.
Main Results:
- The DAPI-DNA complex exhibited specific fluorescence intensity, with RNA contributing less than 2% to the signal.
- The fluorimetric method demonstrated higher sensitivity and provided higher DNA concentration estimates compared to the diphenylamine assay.
- The developed method was less time-consuming than the diphenylamine assay.
Conclusions:
- The fluorimetric method using DAPI offers a sensitive, rapid, and reliable approach for DNA quantification in plant samples.
- This method overcomes the limitations of the diphenylamine assay, providing more accurate DNA concentration data.