Indole acetaldehyde dehydrogenase gene ald2 and overexpression and application
A technology of indoleacetaldehyde dehydrogenase and overexpression, applied in the field of indoleacetaldehyde dehydrogenase gene ald2 and its overexpression and application, can solve the problems of IAA biosynthesis reduction and biosynthesis redundancy
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Embodiment 1
[0033] Embodiment 1 strain activation and preparation
[0034] 1. Observation of strains: NJAU4742 strain is Trichoderma guizhouense, which has the following characteristics: filamentous fungus, hyphae with septal arbuscular shape, conidiophores are smooth, green, the top is enlarged and spherical, and there are small growths on it. Stems produce spores, conidia are smooth spherical, green.
[0035] 2. Culture of strains
[0036] 1) Inoculate NJAU4742 into a solid medium PDA glass petri dish, culture conditions: 28°C, 7 days, wash and scrape the green spores on the petri dish with 5mL sterile water, filter with sterile gauze to sterilize A spore suspension was prepared in a glass bottle for use.
[0037] 2) Inoculate the Trichoderma harzianum NJAU4742 spore liquid into the configured PDA medium according to 1% and shake the flask for culture, the liquid volume is 20%-50% of the volume of the Erlenmeyer glass bottle, and the culture conditions are: temperature is 28°C, 180rmp...
Embodiment 2
[0038] Example 2 The growth-promoting effect of Trichoderma harzianum NJAU4742 on cucumber under hydroponic aseptic conditions
[0039] 1. Growth-promoting effect
[0040] Cucumber seeds used in this study were Lufeng cucumbers from Jiangsu Academy of Agricultural Sciences. Seeds were sterilized in 70% (v / v) ethanol for about 5 minutes, then soaked in 20% sodium hypochlorite (NaClO) for 20 minutes, and then rinsed with sterile water at least 4 times. The sterilized aseptic seeds were germinated in the dark at 30°C for 24 hours, and then the germinated seeds were placed on the beaker column containing the nutrient solution for aseptic culture for 7 days, and the aseptic seedlings with the same growth were transferred into the 50mL container containing 50mL Hoagland's nutrient solution was cultured in a triangular flask, and Trichoderma cells were inserted for interaction experiments. On the 10th day of the interaction, the results of significant growth differences among the tre...
Embodiment 3
[0045] Embodiment 3 Cucumber Root System IAA Content Determination
[0046] Cucumber roots were cut and ground in liquid nitrogen, frozen and ground, dissolved in methanol for extraction, and determined by ELISA method:
[0047] 1) Adding samples: set blank wells, standard wells, and sample wells to be tested respectively. Add 50 μl of sample diluent to the blank well, and add 50 μl of standard substance or sample to be tested in the remaining wells. Be careful not to have air bubbles. Add 50 μl of Detection Reagent A (prepared just before use) to the well, cover the microtiter plate with a membrane, and incubate at 37°C for 1 hour.
[0048] 2) Discard the liquid, shake dry, add 350 μl of 1x Wash Solution, soak for 1-2 minutes, and shake dry. Wash the plate 3 times, put the microplate upside down on absorbent paper for the last time, and blot the liquid.
[0049] 3) Add 100 μl of Detection Reagent B working solution (prepared just before use) to each well, add a membrane, a...
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