Peanut tissue culture method and peanut genetic transformation method
A genetic transformation method and peanut technology, applied in the field of peanut tissue culture method and peanut genetic transformation, can solve the problem that the regeneration system is not suitable for Agrobacterium transformation
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Embodiment 1
[0049] Example 1: Induction of young embryos of peanut variety Shanhua 9 to redifferentiate to form peanut regenerated plants
[0050] (1) Sterilization and soaking of peanut aseptic young embryos: Select fresh young embryos of peanut variety Shanhua No. 9 in the pod expansion stage, carefully peel off the seed embryos containing embryonic leaflets, place them in a 50mL small beaker, add an appropriate amount of 70v / v% alcohol to soak the seeds Embryo for 1min; pour out the alcohol and add an appropriate amount of 0.1wt% HgCl 2 solution, transfer the beaker to a clean bench using HgCl 2 The solution was sterilized for 15 minutes. Rinse with sterilized water 5 times, 5min each time. After rinsing, transfer the seed embryos to a triangular flask, add an appropriate amount of sterile water to seal and soak, and incubate in a 27°C incubator for 16h-18h.
[0051] (2) Acquisition of peanut aseptic explants: in the ultra-clean workbench, cut the embryo leaflet obtained in step (1)...
Embodiment 2
[0056] Example 2: Establishment of regeneration system of peanut varieties with different genotypes
[0057] The specific operation is the same as in Example 1, and the morphology of the bud cluster induction period of different peanut varieties is as follows: figure 2 where a is the lion head enterprise variety, b is the Haihua 1 variety, c is the Luhua 14 variety, d is the Fu peanut variety, e is the Shanhua 9 variety, f is the Fenghua 1 variety, and g is the Weihua variety 8 varieties, h is the four-grain red variety, and i is the Huayu 31 variety.
[0058] The bud cluster induction rate is shown in Table 1: the varieties with the bud cluster induction rate greater than 80% include Shitouqi, Haihua 1, Luhua 14, Fuhuan, Shanhua 9, Fenghua 1, the highest is Shitouqi, reaching 95.36%.
[0059] Table 1 Induction rate of bud clusters of peanut varieties with different genotypes
[0060]
[0061]
[0062] The probability of induced differentiation into seedlings is show...
Embodiment 3
[0066] Example 3: Agrobacterium-mediated genetic transformation of peanut variety Shanhua 9
[0067] (1) Disinfection and soaking of seeds: Select fresh young embryos of peanut variety Shanhua No. 9 in the pod expansion stage, carefully peel off the seed embryos containing embryonic leaflets, add an appropriate amount of 70v / v% alcohol to soak the seed embryos for 1 minute; after the alcohol is drained, add an appropriate amount of 0.1% HgCl 2 The solution was transferred to the ultra-clean workbench for 15 minutes of disinfection; rinsed with sterile water for 5 times, each rinse for 5 minutes. After rinsing, transfer to a conical flask, add appropriate amount of sterile water to seal and soak, and incubate in a 27°C incubator for 16h-18h.
[0068] (2) Pre-cultivation: In the ultra-clean workbench, cut the embryo leaflet obtained in step (1) from the base of the seed embryo, and inoculate it on the bud cluster induction medium. , under the conditions of 16 hours of light an...
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