Praveen K. Saxena

Plantlet formation from isolated protoplasts ofSolanum melongena L.

SummaryMesophyll protoplasts were isolated from axenic shoot cultures ofSolanum melongena by the one-step enzymatic method. Of the different media employed for the culture of protoplasts, a medium modified fromKao andMichayluk (1975) supported sustained mitotic cycles most effectively. Organogenesis from protoplast-derived callus was achieved on transfer toMurashige andSkoogS (1962) medium supplemented with an appropriate auxin and a cytokinin.

Isolation and culture of protoplasts of Capsicum annuum L. and their regeneration into plants flowering in vitro

SummaryAxenic shoot cultures ofCapsicum annuum cv.California Wonder were used as the source for isolation of protoplasts from mesophyll cells. Protoplasts underwent sustained mitotic activity and proliferated to form callus masses on NT or DPD medium enriched with 2,4-D, NAA and BAP each at 1 mg/l level. The callus could be differentiated into whole plants on the differentiation media and plants floweredin vitro under long day conditions.

Differentiation of bud-cells on the protonema of the moss Anoectangium thomsonii. Effect of aspirin and salicylic acid

Summary Differentiation of bud cells on the protonema of the moss Anoectangium thomsonii readily occurs on medium supplemented with phenolic compound-salicylic acid or its acetyl derivative aspirin. Within 20 days of culture the protonema organises to form well-developed gametophytic buds whereas controls, lacking phenolic compounds, remain bud-free.

Development of gametophores from isolated protoplasts of the mossAnoectangium thomsvnii Mitt.

SummaryProtoplasts of the mossAnoectangium thomsonii Mitt. were isolated from preplasmolyzed protonemal filaments, grown in suspension cultures, after the digestion of the cell wall by the enzymes cellulase and macerozyme or driselase. Driselase was more effective than cellulase and macerozyme. After purification these protoplasts were plated in the form of small agar drops in modified Koflers medium without hormones and incubated in the dark at 26 ± 2 °C. Cell walls regenerated within three days and cell divisions started seven days after the initiation of the cultures. When the regenerants were transferred to normal protonemal culture medium and illuminated by 3,000 lux continuous light, a multi-cellular protonema developed which formed leafy gametophores on salicylic acid supplemented medium.

Removal of browning and growth enhancement by polyvinylpolypyrrolidone in protoplast cultures ofCyamopsis tetragonoloba L.

The occurrence of browning in protoplast cultures ofCyamopsis tetragonoloba completely inhibited the growth of protoplast derived colonies. Of the various additives employed to counteract the problem of browning and subsequent necrosis, polyvinylpolypyrrolidone (PVPP) was found most effective. Simultaneous addition of polyvinylpyrrolidone (PVP) to the protoplast culture medium accentuated the effect of PVPP and also improved the frequency of protoplast division.

High frequency regeneration of Funaria hygrometrica protoplasts isolated from low calcium protonemal suspension

Actively growing protonema of Funaria hygrometrica, maintained as suspension in low calcium medium, could be easily transformed into protoplasts employing the enzymes cellulase (3%) or driselase (2%). An increase in the level of calcium for the culture of protonema was inhibitory to the formation of protoplasts. Of the protoplasts plated on modified MS medium, 80% developed cell walls within 36 h and 60–70% divided after 4–5 days. However, if the medium was supplemented with 2,4-dichlorophenoxyacetic acid (2,4-D) and benzylaminopurine (BAP), each at 1 mg/1, the divisions commenced in only 2 days. The regenerated cells on minimal medium produced multicellular and branched protonema which in due course developed leafy gametophytes.

Colony Formation by Cotyledonary Protoplasts of Cyamopsis tetragonoloba L.

Summary Cotyledonary protoplasts of the legume Cyamopsis tetragonoloba resynthesise cell walls and undergo sustained divisions to produce multicellular colonies in two weeks. Preculture of cotyledons in hormone-enriched medium stimulated the division of protoplasts. The division frequency of the protoplasts could also be enhanced considerably by incubation of cotyledons at low temperature of 10°C for 48 h. If the cold conditioning of cotyledons was done in presence of hormones there was a higher frequency of divisions than in individual treatments and the protoplasts divided within 48h of culture. Upon transfer to a medium of low osmotic potential, the colonies developed calli.

Plantlets from mesophyll protoplasts of Solanum xanthocarpum

Young leaves of Solanum xanthocarpum from axenic shoot cultures released viable protoplasts when treated with appropriate enzymes. The protoplasts on culture in modified Murashige and Skoog (1962) medium supplemented with 2,4-dichlorophenoxy-acetic acid (0.5 mg/l), naphtha leneacetic acid (1 mg/l), kinetin (1 mg/l) and organic nutrients of KM (Kao and Michayluk 1975) regenerated to form callus tissue as a result of repeated divisions. Protoplast-derived calli differentiated into shoots on MS medium enriched with kinetin (0.5 mg/l) and rooting could be initiated by transferring the shoot-buds to basal medium.

Optimal conditions for plant regeneration from mesophyll protoplasts of eggplant (Solanum melongena L.)

Abstract Conditions were optimized for the isolation and culture of protoplasts of eggplant (Solanum melongena L.). Leaf tissue obtained from axenic shoot cultures, grown in a controlled environment, was the best source for isolating viable protoplasts. The culture and subsequent regeneration of protoplasts was greatly influenced by cold conditioning of the excised leaves and the composition of the protoplast culture medium. Initial incubation of protoplast cultures in darkness was necessary for inducing sustained cell divisions. More than 70% of the protoplasts divided within 1 week and about 35% of these dividing cells underwent sustained divisions to form larger cell colonies. The protoplast-derived cell colonies developed calli on transfer to a medium of low osmotic potential, and differentiated shoots on a cytokinin-supplemented medium. Regenerated shoots were able to develop into whole plants on a hormone-free culture medium.

Isolation and culture of protoplasts from mesophyll tissue of the legume Cyamopsis tetragonoloba L.

Protoplasts of Cyamopsis tetragonoloba were isolated from leaves of in vitro grown plants. The yield of the protoplasts, their viability and subsequent divisions were greatly influenced by the pH of the media used for isolation and culture of protoplasts. Sustained divisions of the cultured protoplasts were best supported by a modified Kao and Michayluk (1975) nutrient medium containing glucose (0.4 M), NAA (4 mgl−1), 2,4-D (1 mgl−1) and KIN (2 mgl−1 ). The protoplast derived cells developed calli on transfer to Murashige and Skoog (1962) medium supplemented with 1 mgl−1 each of 2,4-D, NAA and KIN.