Regeneration of plants from hypocotyl protoplasts of rapessed (Brassica napus L. var. oleifera) cultivars (original) (raw)
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Rapid and high frequency shoot regeneration from hypocotyl protoplasts of Brassica nigra
Plant Cell, Tissue and Organ Culture, 1993
Protoplasts, isolated from etiolated hypocotyls of seven day old seedlings of Brassica nigra, were cultured in Kao's liquid medium containing 7.2% glucose, 2,4-D (1 mg 1 1), NAA (0.1 mg1-1) and zeatin riboside (0.5 mg 1-1). After initial incubation for 3 days in dark at 25 _+ I°C, cultures were transferred to a photoperiod cycle of 16/8 h and diluted on seventh and tenth day with MS medium containing 3.4% sucrose, 2,4-0 (0.1 mg 1-1) and BAP (1 mg 1 1). About 62% of the cells divided at least once and 46% of them reached 8-16 cell stage in one week. The dividing cell clusters could be plated on agarose medium on the fifteenth day to obtain proliferating minicalli with a plating efficiency of 1.8%. 56.8% of minicalli, regenerated shoots on a regeneration medium containing 2 IP and IAA at 1 and 0.2 mg 1 1 respectively. The in vitro produced shoots were rooted in MS medium containing 1 mg 1 1 IBA and established in soil without difficulty. The time taken for protoplasts to develop into plants varied from 9 to 10 weeks.
A simple culture method for Brassica hypototyl protoplasts
Plant Cell Reports, 1985
Hypoco~,l protoplasts from oil rape, Brassica napus L. cv. Isuzu were cultured in the d a r k-~ in a modified Nitsch and Nitsch medium containing 13% sucrose, 5 g/l Ficoll, 0.5 mg/l BAP, l mg/l NAA and 0.5 mg/l 2-4 D. Protoplasts floated on the surface of the medium and developed into microcolonies 0.5 mm in diameter in 4-6 weeks. The microcolonies also remained on the surface of the medium. Transfer to MS medium supplemented with 200 mg/l casein hydrolysate, 5mg/l BAP, 0.5 mg/l NAA and solidified with 0.6% agarose induced shoot regeneration in 3-4 weeks.
Agricultural and Food Science
Protoplast cultures were prepared from 6-day-old hypocotyls of six spring, seven winter cultivars of Brassica napus L. and one line of Brassica campestris L. The molarity of enzyme solution was raised to 0,714 M mannitol resulting in well manipulable, cytoplasm dense protoplasts. In the protoplast purification procedure density gradient centrifugation was used to minimize physical damage of protoplasts. Three different protoplast culture systems —(1) liquid, (2) 2nd day embedded, (3) directly embedded in low melting agarose were compared. The two different protoplast embedding techniques resulted in the same efficiency of cell division as the liquid culture method and over this fact the colony browning was avoided. Using protoplast agarose-embedding and culture techniques, healthy calli were obtained for plant regeneration experiments. Incorporation of silver nitrate into the regeneration medium improved the efficiency of plant regeneration in responsive genotypes and the regenerati...
Protoplast culture and regeneration from Brassica oleracea 'rapid cycling'and other varieties
Plant cell, tissue and organ culture, 1993
The results are reported for the first time on successful plant regeneration from mesophyll-derived protoplasts of 'rapid cycling' B. oleracea. Comparative data were also presented on plant regeneration from mesophyll-derived protoplasts of two other varieties namely var. botrytis and var. gemmifera. It was found that a modified Pelletier (Pelletier et al. 1983) protocol is highly beneficial for protoplast culture and plant regeneration from mesophyll-derived protoplasts. The plating efficiency of B. oleracea 'rapid cycling' protoplasts was, in the best combination of isolation method, culture technique and culture media 4.5% + 0.4% and the plant regeneration frequency approximately 15%. Plant regeneration was further improved by transferring the calli from the D medium of Pelletier to a callus growth medium (MSll) and subsequently to the K 3 regeneration medium of Glimelius (Glimelius 1984). Various factors influencing plating efficiency and plant regeneration from mesophyll protoplasts of B. oleraeea are discussed.
Protoplast culture and plant regeneration from Brassica carinata Braun
1987
Protoplasts isolated from hvDocotvls of three-day-old seedlinqs of Brassica carinBta (Rraun) cv R-212~ were cultured in a modified Nitsch and Nitsch l i n u i d medium containinQ 13% sucrose, 0.4% F i c o l l , 0.25 mq/l BA, 0.5 mq/l NAA and Q.5 mg/l 2,4-D. The densit~ of medium caused the DrotoDlasts and the developinn microcalli to f l o a t on the surface of the l i n u i d medium whereas a l l debris and lysed c e l l s sank to the bottom of the culture plate. After 4-6 weeks developing microcalli were approximately O. 5 mm in diameter and were transferred onto MS medium containinq 3% sucrose, ~.4% agarose, 200 m~/l casein h~drolysate~ 5 mq/l ~A and ~.5 mq/l NAA, pH 5.7. Approximately 20% of the c a l l i transferred to this medium produced plantlets.
Plant regeneration from root protoplasts of Brassica
Plant Science Letters, 1982
Protoplasts isolated enzymatically from roots of Brassica alba (White Mustard), B. campestris {Turnip), B. napus (Rape) and B. oleracea (Cabbage), divided to form callus. Plant regeneration was obtained from protoplast derived tissues of B. napus and B. oleracea, but only rhizogenesis was observed with B. campestris. Tissues of B. alba remained undifferentiated. The suitability of root protoplasts for genetic manipulations in the genus Brassica is discussed. *On leave from the Institute of Plant Physiology, Academia Sinica, Shanghai, China. Abbreviations: BAP, 6-benzylaminopurine; CPA, p-chlorophenoxyacetic acid; IAA, 3-indoleacetic acid; NAA, a-naphthaleneacetic acid; 2,4-D, 2,4-dichlorophenoxyacetic acid. 0304--4211/82/0000--0000/$02.75