Method for establishing system for inducing and regenerating sisal hemp stem tip calluses
A technology of callus and shoot tip, which is applied in the field of tissue culture and seedling cultivation, can solve the problems of slow proliferation of callus, vitrification propagation coefficient, and low propagation coefficient, and achieves easy implementation steps, loose implementation conditions, and simple implementation steps Effect
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Embodiment 1
[0031]Take the shoot tips of sterile shoots kept in our laboratory as explants, cut them longitudinally and inoculate them in callus induction medium (MS﹢NAA 0.1-1.0mg / L﹢6-BA 1.5-3.5mg / L) Culture in medium dark for 10 days, peel off the outer leaves and transfer to semi-light conditions for culture. After the callus is induced, take light yellow-green spherical or block callus and inoculate it on the callus subculture medium (improved SH2+ 6-BA 1.5-4.0mg / L+NAA 0.05-0.5mg / L+IBA 0.05-0.5mg / L) for subculture, and after green buds appeared on the surface of the callus, it was transferred to the differentiation medium ( Improved SH1+6-BA 1.0-5.0mg / L+NAA 0-1.5mg / L+IBA 0-1.5mg / L) to induce plant regeneration. The culture conditions are: pH 5.8-6.0, canal gum 6.8-8.0 g / l, sucrose 30g / l, light 12-14h, dark 10-12h, temperature 28±2°C; a total of 58 explants were inoculated this time. 1021 regenerated plants were obtained, the callus induction rate was 93.6%, and the differentiation rat...
Embodiment 2
[0033] The stem tips of H.11648 hemp sucking buds were used as the material for disinfection. The roots and some old leaves were removed first, and the remaining leaves were peeled off with a scalpel blade under running water, and treated with 0.3% potassium permanganate solution for 30 minutes, and then the Soak the clean bench with 75% alcohol for 1 min, then soak with 0.1% mercuric chloride solution for 15-30 min, and finally rinse with sterile water for 3-5 times. Dry and cut off part of the explants, inoculate them on the improved SH1+6-BA1.0-5.0mg / L medium after longitudinal cutting, the culture conditions are 12-14 hours of light per day, 10-12 hours of dark culture, light intensity 2000lux, culture temperature is 28±2℃. After the cluster buds grow out, use the tips of the vigorous cluster buds as explants, cut them longitudinally and inoculate them in callus induction medium MS﹢6-BA 1.5-3.5mg / L﹢NAA 0.1-1.0mg / L Culture in dark for 10 days, peel off the outer leaves and...
Embodiment 3
[0035] Disinfect the stem tips of sisal H.11648 10-15cm bulbil seedlings, peel off the leaves with a scalpel under running water, treat with 0.3% potassium permanganate solution for 30 minutes, and then use 75% alcohol on the ultra-clean workbench Soak for 1 min, then soak in 0.1% mercuric chloride solution for 15-30 min, and finally rinse with sterile water for 3-5 times. Dry and cut off part of the explants, inoculate them on the improved SH1+6-BA1.0-5.0mg / L medium after longitudinal cutting, the culture conditions are 12-14 hours of light per day, 10-12 hours of dark culture, light intensity 2000lux, culture temperature is 28±2℃. After the clustered buds are induced, take the shoot tip of the vigorous sterile seedling, cut it longitudinally, inoculate it in the callus induction medium and culture it in the dark for 10 days, peel off the outer layer of leaves and transfer it to half-light condition for cultivation, and take a light yellow green seedling. Spherical or ma...
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