Plant Tissue Cult. & Biotech. 24(1): 93-100, 2014 (June)

Isolation of Protoplasts from Nepenthes - A Plant Carnivore

Jaylen Sweat and Michael S. Bodri

Department of Biology, University of North Georgia, Dahlonega, GA 30597, USA

Key words: Cellulase, In vitro culture, Nepenthes, Protoplast

Abastract

Protoplasts were isolated from the lamina of greenhouse grown Nepenthes ampullaria and the hybrid N. ?Rokko? Exotica in order to develop a protocol for protoplast isolation suitable for wild species of Nepenthes. Various molarities utilizing mannitol or sorbitol and different enzyme mixtures and concentrations as well as incubation times were evaluated to maximize protoplast yield and viability. The most effective treatment, a 4 hrs incubation at 40 rpm and 25^?C in a solution consisting of 0.5 M sorbitol, 5% cellulase ?Onozuka? R-10, 0.5% macerozyme R-10, and 0.3% pectolyase Y-23, generated 4.35 ? 10⁶ protoplasts/ gfw of which 62.1% were viable. Culture was attempted in respect of regeneration of the cell wall, however, no cell division was observed.

Download Full Article