Cryopreservation and plant regeneration method of vegetative propagation flower narcissus
A technology of ultra-low temperature preservation and vegetative reproduction, applied in the fields of plant regeneration, botanical equipment and methods, horticultural methods, etc., can solve the problems of loss of ornamental value and commerciality, degeneration of species, degeneration, etc., and achieve the simple and feasible method of plant regeneration. , restore good growth condition
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Embodiment 1
[0022] Example 1, Induction of Small Bulbs of Chongming Narcissus
[0023] Such as figure 2 Shown and illustrated is the flow chart of the Chongming narcissus small bulb induction operation flow chart of the present invention. The operation steps are: a', select 3-4 year old Chongming narcissus bulbs; b', sterilize the bulbs of step a'with 70% alcohol 1 min, sterilize with 0.1% mercury for 10-20 min, then rinse with sterile water; c', inoculate the bulb cut pieces of step b'with 1 mg·L -1 BA and 1 mg·L -1 Small bulbs were induced on NAA's MS medium, where the induction culture temperature on the medium was 25±2℃, and the light was 12 h·d -1 ,Light intensity 36 μmol m -2 s -1 , After 3 months of induction, small bulbs with a diameter of 0.5 to 1 cm are obtained.
Embodiment 2
[0024] Example 2, Chongming narcissus pre-training
[0025] Such as figure 1 As shown, the description is the operation flow chart of the preservation method of the present invention. The operation method of the preservation method is: a. Induce the bulbs of vegetatively propagated flower narcissus, and cut the stem tips of 2~5mm in size; b. The shoot tips of root primordium and bud primordium are pre-cultured, and then placed in the pretreatment solution for 15-30 minutes at room temperature; c. Transfer the shoot tips treated in the pretreatment solution to the vitrification protectant for ice bath Treat for 60-120min; d. Transfer the ice-bath treated stem tip to a freezing tube containing fresh pre-cooled vitrification protectant; e. Put the above freezing tube into liquid nitrogen quickly for storage.
Embodiment 3
[0026] Example 3. The effect of stem tip size on the survival rate of Chongming Narcissus under cryopreservation
[0027] The sucrose concentration of 1~1.5 mm, 2~3 mm, 4~5 mm stem tips is 0.3 mol·L -1 After pre-cultured in MS medium for 5 days, it was treated with pretreatment solution for 20 min, and then treated with vitrification protectant ice bath for 60-120 min. The preferred treatment time was 80 min. The result of resuming culture after freezing storage showed: The survival rate of 2~3mm diameter is the highest, up to 80%. The survival rate of stem tips less than 2 mm is only 40%, and the survival rate of stem tips greater than 4 mm is only 10%.
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