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Interspecies protoplast fusion in Larix: Comparison of electric and chemical methods

Interspecies protoplast fusion in Larix: Comparison of electric and chemical methods

In Vitro Cellular & Developmental Biology Plant 34(3): 212-217, July-Sept

Larix was chosen for the study on interspecies protoplast fusion due to its ability to regenerate plants from protoplasts derived from embryogenic cultures. L. laricina line L2 was used in fusion experiments with either L. X eurolepis line L6 or L. X leptoeuropaea line L5. A method of unambiguous labeling of parental protoplasts prior to fusion was developed using vital fluorescent dyes. Of a number of dyes tested, only rhodamine B hexyl ester chloride (R6) and 3,3'-dihexyloxcarbocyanine iodide (DiOC6) stained the protoplasts in a consistent and uniform fashion. The fusion of mixed parental protoplasts that were internally labeled was carried out either in the presence of a 20% polyethylene glycol (PEG) solution or in an electric field. The progress of fusion was readily observed, taking only minutes under the experimental conditions. The fusion products could be identified by dual fluorescence several h after the onset of fusion. Heterofusion frequencies of approximately 18% and 6% in the presence of PEG and an electric field, respectively, were attained. Postfusion cultures between L. X laricina protoplasts and protoplasts of L. X leptoeuropaea gave rise, to cell colonies and between L. laricina and L. X eurolepis, to mature somatic embryos.

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Accession: 008901996

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DOI: 10.2307/20064985

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