Experiments on the induction of α-amylase formation by gibberellin
Gibberellins are produced in the dextrinous layer of the embryo after water absorption and germination of barley or wheat seeds. These gibberellins, when released into the endosperm, can induce and increase the activity of some hydrolytic enzymes such as β-1,3-glycosidase, protease, ribonuclease, α-amylase, etc. The most thorough study is on the effect of gibberellins on α-amylase in the endosperm of barley. One of the most thorough and in-depth research is the formation of α-amylase induced by gibberellin in barley flour layer. Gibberellin induces or enhances the activity of some hydrolytic enzymes which are important for material transformation and organogenesis during seed germination. In seeds without embryo, there is no α-amylase in the endosperm because gibberellin cannot be produced.
Operation method
Experiments on the induced formation of alpha-amylase by gibberellin
Principle
Gibberellins are produced in the dextrinous layer of the embryo after water absorption and germination of barley or wheat seeds. These gibberellins, when released into the endosperm, induce and increase the activity of some hydrolytic enzymes such as β-1,3-glycosidase, protease, ribonuclease, α-amylase and other enzymes. Among them, the most thoroughly and deeply researched is the formation of α-amylase induced by gibberellin in the barley dextrin layer. Gibberellin induces or enhances the activities of some hydrolytic enzymes which are important for material transformation and organogenesis during seed germination. Seeds without embryos have no α-amylase in the endosperm because they cannot produce gibberellin. Move I. Materials and equipment For more product details, please visit Aladdin Scientific website.
Wheat seeds
Spectrophotometer or photoelectric colorimeter, beakers, pipettes, water bath, test tubes, razor blades, tweezers, penicillin vials, thermostat.
Medicines
0.1% starch solution: 1 g of soluble starch plus 50 ml of distilled water, boiling water bath until the starch is completely dissolved, then add KH2PO4 8.16 g, to be dissolved and then fixed to 1000 ml.
2×10-5 M GA3 solution: 680 mg of GA3 was dissolved in a small amount of 95% ethanol and then concentrated to 1000 ml.
I2-KI solution: 0.600 g of potassium iodide and 0.060 g of iodine (I2) were dissolved in a small amount of 0.05 N HCl and mixed, and then volumes were set to 1000 ml with 0.05 N HCl.
1% sodium hypochlorite solution.
10-3 M Acetic acid buffer: 590 ml of 10-3 M sodium acetate solution was mixed with 410 ml of 10-3 M acetic acid solution, 1 g of streptomycin was added and shaken well.
II. Experimental steps
1. Select 50 barley seeds with full grains of uniform size, cut each seed into two halves with and without embryo with a razor blade, and put them into the newly prepared 1% sodium hypochlorite solution to sterilize for 15 minutes, and then take them out and rinse them with sterile water for several times. The seeds were sucked and swelled for 48 hours under aseptic conditions.
2. 2×10-5 M GA3 was diluted into 2×10-6 M , 2×10-7 M , 2×10-8 M solutions; 10-3 M acetic acid buffer was diluted 1 times.
3. 6 vials of penicillin were taken, numbered, and reagents and materials were added according to the table below: 
4．Put the vials in a thermostat and incubate them at 25°C with oscillation for 24 hours. If there is no oscillator, it should be shaken frequently.
5. Determination of amylase activity: Pipette 0.2 ml of supernatant from each vial into a test tube containing 1.8 ml /0.1% starch solution of the corresponding label and mix well. Then keep warm in a water bath at 30℃ for exactly 10 minutes (the holding time is best determined by preparatory experiments, and a reaction time with an optical density of 0.4-0.6 is preferred). Then add 2 ml of I2-KI solution, dilute to 5 ml with distilled water and shake well. This solution is blue and can be colorimetrically compared with a red filter or at a wavelength of 580n M. The color of the solution can be compared with a distilled water as a control. Distilled water was used as a control.