Plant stress tolerance-associated protein GUI as well as coding gene and application thereof
A protein and transgenic technology, applied in the fields of plant peptides, biochemical equipment and methods, microorganisms, etc., can solve problems such as the role report has just started, and achieve the effect of improving drought tolerance.
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Embodiment 1
[0033] Embodiment 1, gui improve the drought resistance of plants
[0034] 1. Acquisition of gui mutants
[0035] The T-DNA insertion mutants of the GUI gene were purchased from the SALK library and the NASC library respectively, in which the T-DNA of SALK 092071 was inserted in the second exon, and the N335814 was inserted in the fourth exon, both of which were the genes of the gene. knockout mutants.
[0036] 2. Identification of drought tolerance of transgenic plants
[0037] The drought tolerance of Arabidopsis WT (Col-0) and gui homozygous mutants (60 strains each) were identified. The experiment was repeated three times, and the results were averaged.
[0038] The normal growing seedlings germinated for 15 days were not watered until the wild-type plants withered (about 2 weeks), and then rewatered for a week, the phenotype was observed, photographed and the survival rate was counted. see photos Image 6 . The survival rate of Arabidopsis WT was 10%, but almost 100...
Embodiment 2
[0040] Embodiment 2, the expression characteristic of semi-quantitative PCR analysis GUI
[0041] 1. Coercion treatment
[0042] Take soybean Tiefeng No. 8 seedlings grown at room temperature for about 20 days and carry out the following treatments:
[0043] For drought treatment, Arabidopsis WT grown on MS medium for 8 days was taken out and placed on filter paper soaked with PEG solution. After 12 hours of cultivation, the material was taken out, quickly frozen with liquid nitrogen, and stored at -80°C for later use. The salting treatment is to take out the Arabidopsis WT grown on MS medium for 8 days and place it on MS medium supplemented with 200mM NaCl. After 12 hours of light culture, take out the materials respectively, freeze them with liquid nitrogen, and store them at -80°C for later use. . The abscisic acid treatment is to take out Arabidopsis WT grown on MS medium for 8 days and place them on MS medium supplemented with 100 μM abscisic acid (ABA). Save for later...
Embodiment 3
[0051] Example 3, GmNF-YB1 subcellular localization analysis
[0052] 1. Construction of recombinant expression vector
[0053] 1. Cloning of GUI gene
[0054] Primer pairs (GUI-F and GUI-R) were designed according to the sequence of the GUI gene, Sal I and BamH I restriction sites were introduced at the ends of the primers, and GUI was amplified by PCR using the cDNA of Arabidopsis wild-type Col-0 as a template Gene.
[0055] GUIF: 5'-GCGTCGACATGGCAGCGTTAAGCCAC-3';
[0056] GUIR: 5'-CGGGATCCTCAAGGTGATCTTGAGACTAAAC-3'.
[0057] The PCR amplification product was subjected to 1% agarose gel electrophoresis, and a band of about 1.4 Kb was recovered and purified using Agarose Gel DNA Purification Kit Ver.2.0 (TaKaRa Company, Code No.: DV807A).
[0058] 2. Construction of recombinant expression vector
[0059] ① Use restriction endonucleases Sal I and BamH I to digest step 1 to recover the purified PCR product, and recover the digested product;
[0060] ② Digest pEZR(K)-LC (a...
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