Wolfberry anther culture breeding differential medium

[0034] Example 1

[0035] Prepare the culture medium with the following formula:

[0036] Macroelements: KNO 3 950mg/L, NH 4 NO 3 825mg/L, KH 2 PO 4 675mg/L, MgSO 4 ∙7H 2 O 1500mg/L, Glutamine 390mg/L, CaCl 2 ∙2H 2 O 220mg/L;

[0037] Trace elements: MnSO 4 ∙4H 2 O 18mg/L, ZnSO 4 ∙7H 2 O 27.5mg/L, H 3 BO 3 6.25mg/L, KI 0.8mg/L;

[0038] Iron salt and complexing agent: FeSO4 ∙7H 2 O 42.5mg/L, Na 2 -EDTA∙2H 2 O 56mg/L;

[0039] Organic ingredients: inositol 50mg/L, vitamin B1 0.4mg/L, vitamin B6 0.5mg/L, niacin 5.0mg/L, adenine 22.5mg/L, proline 2.0mg/L, folic acid 0.4mg/L , Biotin 0.1mg/L;

[0040] Inorganic additives: activated carbon 9g/L, N-methyl-nitrosourea 27.5mg/L;

[0041] Plant growth regulator: 2,4-D 1.0mg/L, KT 1.7mg/L;

[0042] Physiologically active substances: Lycium barbarum root extract 12.5g/L;

[0043] Carbon source: sucrose 27.5g/L, fructose 13.5g/L;

[0044] Coagulant and other additives: plant gel 5.75g/L, sorbitol 22.5g/L, hydrolyzed protein 1.0g/L.

[0045] Lycium barbarum varieties Ningqi No. 3 and Lycium barbarum Huangguo were selected as anther donors for anther culture, and flower buds in the middle and late stage of uninucleate development were taken from the two varieties of Lycium barbarum plants that were robust and free of diseases and insect pests at the early flowering stage or full flowering stage. As a test material, callus of Lycium barbarum was induced. After the callus is formed (when the diameter reaches about 2 mm), the Lycium barbarum callus is inoculated into the differentiation medium. After the Lycium barbarum callus completely differentiates into Lycium barbarum seedlings, the differentiation rate of the Lycium barbarum callus green shoots and the albino seedling rate are counted.

[0046] Green seedling differentiation rate (%) = number of differentiated green seedlings/number of transferred callus × 100%; rate of albino seedlings (%) = number of albino seedlings/number of transferred callus × 100%.

[0047] Example 2

[0048] Prepare the medium with the following formula (optimized screening of carbon source combination):

[0049] Macroelements: KNO 3 950mg/L, NH 4 NO 3 825mg/L, KH 2 PO 4 675mg/L, MgSO 4 ∙7H 2 O 1500mg/L, Glutamine 390mg/L, CaCl 2 ∙2H 2 O 220mg/L;

[0050] Trace elements: MnSO 4 ∙4H 2 O 18mg/L, ZnSO 4 ∙7H 2 O 27.5mg/L, H 3 BO 3 6.25mg/L, KI 0.8mg/L;

[0051] Iron salt and complexing agent: FeSO 4 ∙7H 2 O 42.5mg/L, Na 2 -EDTA∙2H 2 O 56mg/L;

[0052] Organic ingredients: inositol 50mg/L, vitamin B1 0.4mg/L, vitamin B6 0.5mg/L, niacin 5.0mg/L, adenine 22.5mg/L, proline 2.0mg/L, folic acid 0.4mg/L , Biotin 0.1mg/L;

[0053] Inorganic additives: activated carbon 9g/L, N-methyl-nitrosourea 27.5mg/L;

[0054] Plant growth regulator: 2,4-D 1.0mg/L, KT 1.7mg/L;

[0055] Physiologically active substances: Lycium barbarum root extract 12.5g/L;

[0056] Coagulant and other additives: plant gel 5.75g/L, sorbitol 22.5g/L, hydrolyzed protein 1.0g/L.

[0057] Set the following 6 carbon source combinations: sucrose 40g/L; sucrose 35g/L, fructose 5g/L; sucrose 30g/L, fructose 10g/L; 30g/L; fructose 40g/L.

[0058] The two Lycium barbarum varieties were also selected as anther donors, the operation method and cultivation method were the same as in Example 1, and the differentiation rate and albino seedling rate of Lycium barbarum callus green shoots were counted.

[0059] Example 3

[0060] Prepare the medium with the following formula (optimized screening of coagulants):

[0061] Macroelements: KNO 3 950mg/L, NH 4 NO 3 825mg/L, KH 2 PO 4 675mg/L, MgSO 4 ∙7H 2 O 1500mg/L, Glutamine 390mg/L, CaCl 2 ∙2H 2 O 220mg/L;

[0062] Trace elements: MnSO 4 ∙4H 2 O 18mg/L, ZnSO 4 ∙7H 2 O 27.5mg/L, H 3 BO 3 6.25mg/L, KI 0.8mg/L;

[0063] Iron salt and complexing agent: FeSO 4 ∙7H 2 O 42.5mg/L, Na 2 -EDTA∙2H 2 O 56mg/L;

[0064] Organic ingredients: inositol 50mg/L, vitamin B1 0.4mg/L, vitamin B6 0.5mg/L, niacin 5.0mg/L, adenine 22.5mg/L, proline 2.0mg/L, folic acid 0.4mg/L , Biotin 0.1mg/L;

[0065] Inorganic additives: activated carbon 9g/L, N-methyl-nitrosourea 27.5mg/L;

[0066] Plant growth regulator: 2,4-D 1.0mg/L, KT 1.7mg/L;

[0067] Physiologically active substances: Lycium barbarum root extract 12.5g/L;

[0068] Carbon source: sucrose 27.5g/L, fructose 13.5g/L;

[0069] Other additives: sorbitol 22.5g/L, hydrolyzed protein 1.0g/L.

[0070] The addition combination of coagulant was set to the following three types: plant gel 1.9g/L, agar 4.7g/L; plant gel 3.8g/L, agar 2.3g/L; agar 7g/L.

[0071] The two Lycium barbarum varieties were also selected as anther donors, the operation method and cultivation method were the same as in Example 1, and the differentiation rate and albino seedling rate of Lycium barbarum callus green shoots were counted.