Zachrisson, A, and Bornman, C H, 1984, Application of electric field fusion in plant tissue culture, -Physiol, Plant, (il: 314-320, Manipulation of protoplasts via fusion and organcllc transfer is expected to be facilitated by the technique known as electric field fusion. Construction and tise ate described of three flow-through fusion chambers that incorporate flat-sided electrodes in a manner that makes fusion of ptotoplasts possible throughout the cltantbcrs' total volume (4, 49 or 110 [i\) under constant electrical, chemical and physical conditions, Brassica napus L, protoplasts subjected to Itisogenic conditions, that is, application of voltages that induce reversible membrane bteakdown, were capable not only of survival but also of cell wall tcsynthcsis, cell division and subscqttcnt growth and development, Intraspecific (B. napus x B. napus), interspecific (B. napus X B. campestris L.) and intcrgcncric (B. ini/nis x Primula acaulis L,) fusion and engullment events were followed by using on the one hand autolluoresccncc and fluorescein isothiocyanate as respective markers or on the other hand autolluorcsccnce and vacuolar anthocyanin (Primula). Properties and merits of flat-sided veisus cylindrical electrodes are discussed.

The current status of electromanipulation, that is, electrofusion and electroporation, of plant protoplasts is reviewed. Parameters for electromanipulation as well as their practical implications are discussed. Some comparisons with the use of polyethylene glycol are made and the advantages of electromanipulation are considered.

Datura innoxia cell suspension-derived protoplasts were anchored to Cytodex 1 microcarriers pre-swollen in buffered concanavalin A. As many as 34 protoplasts were estimated to attach per microcarrier, in comparison to a potential 47 as determined from a model based on random anchorage. Fluorescein diacetate was used as localizing agent as well as to assess viability. When included in the swelling medium fluorescence was observed almost instantaneously, first in the protoplast at its interface with the microcarrier, and later throughout the cytoplasm. However, the dye was not conjugated with the lectin, and leakage eventually resulted in fluorescence also of non-anchored protoplasts. Fluorescein-labelled concanavalin A on the other hand permitted detection of microcarriers but not of anchored protoplasts, suggesting the use of differentially fluorescing microcarriers, as an aid in identification of fusion partners.

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