Phytophthora cinnamomum effector protein Avh49 and application thereof
An effector protein, avh49 technology, applied in applications, recombinant DNA technology, angiosperms/flowering plants, etc., can solve the problems of different effector protein structures, and achieve the effect of reducing plant mortality and significant application value
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Embodiment 1
[0049] Example 1 Acquisition of the gene encoding effector protein Avh49 of Phytophthora camphora
[0050] The Phytophthora camphora strain was selected as the test material, and the gene sequence of the effector gene Avh49 was obtained according to the whole genome sequence of Phytophthora camphora. Then design primers p1 and p2 according to the obtained gene fragments, and amplify and screen the obtained target gene fragments.
[0051] 1. Using CTAB-SDS method to extract high-quality Phytophthora camphora genomic DNA
[0052] Cultivate Phytophthora camphora strains on solid V8 medium plate (recipe: 340ml V8 fruit juice plus 3.4g calcium carbonate, mix well with glass rod, filter the liquid with two layers of filter cloth, add nine times the volume of pure water to dilute, take 200ml in 250ml Add 3g of agar powder to the Erlenmeyer flask, and sterilize at 121°C for 20 minutes for later use.) After culturing in a constant temperature incubator at 25°C for 3 days, take 8-12 pi...
Embodiment 2
[0065] Example 2 Homology analysis between different effector proteins
[0066] According to the analysis of the genome information of Phytophthora camphora, the amino acid sequence encoded by Av h49 cloned in this study and Phytophthora camphora (P.cinnamomi), P.cinnamomi var.robiniae and Phytophthora camphora The comparison of other effector molecules of P. cinnamomi var. parvispora showed that the homology between PcinnAvh49 and other effector proteins was not high, such as figure 2 As shown, it can be seen that there is no research reference between effector proteins, and bioinformatics analysis and functional experimental verification of gene proteins are needed to determine the function of each effector protein.
Embodiment 3
[0067] Example 3 Analysis of the expression pattern of Avh49 gene during the infection of apple by Phytophthora camphora
[0068] 1. Phytophthora camphora strains infecting apples
[0069] The mycelium pieces stored in the refrigerator at 4°C were activated on V8 solid medium, and cultured at 25°C for 3 days. Place a piece of sterilized filter paper of appropriate size on the bottom of the sterilized Petri dish and moisten it with sterilized water. Wipe the surface of the apple with an alcohol swab. Use a scalpel to cut small squares about 1 cm2 and 1 cm thick on three sides of the apple to remove the pulp. Pick out the mushroom cake and stuff it into the incision on the surface of the apple, only stuffing two sides, and one side as a blank control of the solid medium. Plug the incision with absorbent cotton and drip with sterile tap water. Pack it into a constant temperature incubator with a tray and cultivate it. After infecting for 24h, 48h, 72h, and 96h respectively, ...
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