Application of atga2ox1 gene in regulating stress resistance of plants and method for cultivating stress-resistant plants
A plant stress resistance and stress resistance technology, applied in the field of plant bioengineering, can solve problems such as reduced survival rate, and achieve the effects of high success rate, simple operation, and improved stress resistance ability
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Embodiment 1
[0071] Embodiment 1 Plant expression vector construction and genetic transformation of maize
[0072] Clone the full-length cDNA of the AtGA2ox1 gene (GENE BANK accession number: AT1g78440), connect it into the monocotyledonous plant expression vector pCAM-UGN, obtain the expression vector containing the AtGA2ox1 gene, and name it pCAM-UGN-GA2ox, such as figure 1 shown. Agrobacterium-mediated method was used to infect HiII maize immature embryos to obtain transgenic maize plants.
Embodiment 2
[0073] The identification of embodiment 2 transgenic plants and the expression analysis of exogenous gene
[0074] Identification of transgenic plants:
[0075] Total DNA was extracted from leaves of transgenic plants by CTAB method. Design PCR amplification primers according to the AtGA2ox1 gene sequence, as shown in the following table:
[0076] name Numbering Primer sequence (5'-3') AtGA2ox-445F SEQ ID NO.1 GGTTCGGGTCCACTATTT AtGA2ox-445R SEQ ID NO.2 CTATGCCTCACGCTCTTG
[0077] The PCR reaction system is:
[0078] Reagent Volume (μL) Taq Buffer(10×) 2.0 dNTP (2.5mM) 2.0 AtGA2ox-445F (10μM) 1.5 AtGA2ox-445R (10μM) 1.5 Taq DNA polymerase 0.2 wxya 2 o
to 20.0
[0079] The PCR reaction conditions are:
[0080]
[0081] The amplified product was subjected to 1% agarose gel electrophoresis, and the PCR electrophoresis result was observed under an ultraviolet gel imager.
[0082] E...
Embodiment 3
[0086] Phenotypic Analysis of Example 3 Corn Plant Drought Stress
[0087] Choose corn plants grown in a greenhouse at a temperature of 27°C±2°C, potted (10×10cm), peat soil: vermiculite (5:1, v / v), and grown for 21 days under natural light conditions (each genotype Each 10 strains) were subjected to drought stress treatment, and watering was stopped for 9 days. The maize leaves on the 9th day after drought stress were taken, frozen in liquid nitrogen, and stored in a -80°C refrigerator for subsequent determination of relevant physiological indicators. The amount of substrate and moisture used for each independent experimental material was kept the same.
[0088] In this example, to further evaluate whether the overexpression of the AtGA2ox1 gene can improve the drought stress resistance of maize plants, three independent transgenic maize lines (1#, 2#, 4#) were randomly selected for relevant experimental analysis. Corn plants at the seedling stage of 21 days were selected, ...
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