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A rapid, simple method for nuclei isolation from plant protoplasts.

K Ohyama¹, L E Pelcher, D Horn

¹National Research Council of Canada, Prairie Regional Laboratory, Saskatoon, Saskatchewan S7N 0W9.

Plant Physiology

|August 1, 1977

Summary

Researchers developed a quick method to isolate soybean nuclei. This technique successfully separated nuclear DNA from mitochondrial and chloroplast DNA, aiding plant cell research.

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Area of Science:

Plant molecular biology
Cell biology
Biochemistry

Background:

Soybean (Glycine max L. Merr.) is a vital crop.
Efficient isolation of plant cell components is crucial for molecular studies.
Distinguishing nuclear material from other organelles is a common challenge.

Purpose of the Study:

To develop a rapid and simple method for isolating and purifying nuclei from soybean protoplasts.
To characterize the biochemical activity of isolated soybean nuclei.
To differentiate nuclear DNA from mitochondrial and chloroplast DNA in soybean.

Main Methods:

Isolation and purification of nuclei from soybean protoplasts.
Assay of amino acid and RNA synthesis in isolated nuclei.

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Analysis of DNA by Cesium Chloride (CsCl) density gradient centrifugation.

Main Results:

The method provided rapid and simple isolation of soybean nuclei.
Isolated nuclei showed active amino acid incorporation and RNA synthesis.
DNA synthesis was not detected in the isolated nuclei.
CsCl density gradient centrifugation revealed a single DNA band from nuclei, contrasting with three bands (nuclear, mitochondrial, chloroplast DNA) from protoplasts.

Conclusions:

The described method effectively isolates pure soybean nuclei.
The isolated nuclei are transcriptionally active.
This technique allows for the clear separation of nuclear DNA from organellar DNA in soybean.