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Plant and Cell Physiology, 1995, Vol. 36, No. 8 1493-1502
© 1995

Aluminum-tolerant Protoplasts from Roots Can be Collected with Positively Charged Silica Microbeads: a Method Based on Differences in Surface Negativity

Tadao Wagatsuma, Kouichi Jujo, Satoru Ishikawa and Takumi Nakashima

Laboratory of Plant Nutrition and Soil Science, Faculty of Agriculture, Yamagata University Tsuruoka 1-23, Yamagata, 997 Japan

A new technique was developed for the collection of Al-tolerant plant cells. The procedure takes advantage of the difference in surface charge between Al-tolerant and Al-sensitive protoplasts. Equal volumes of freshly prepared, positively charged silica microbeads (0.05% w/v) and purified protoplasts derived from root tips (2x105 ml-1) of rice, maize or pea were mixed and then centrifuged on a discontinuous Ficoll gradient. Intact protoplasts from the Al-tolerant plant were recovered mostly in the bottom fraction, those from the Al-sensitive plant were recoverd at the uppermost interface, and those from the plant with intermediate tolerance to Al were collected at the middle interface. Moreover, the pattern of fractionation was unrelated to the diameter of the protoplasts. The technique should be useful in future investigations of the physiological and biochemical properties of Al-tolerant cells or their plasma membranes.

(Received April 19, 1995; Accepted September 1, 1995)
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