Application of Proteins Related to Rice Grain Shape and Leaf Angle and Their Encoding Genes
A technology of leaf angle and grain type, applied in the field of genetic engineering, can solve problems such as large heritability
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Embodiment 1
[0040] Example 1. Phenotypic analysis of rice grain shape and leaf angle related mutant slg-D
[0041] The mutant slg-D associated with granulotype and leaf angle was screened in a library of T-DNA insertion mutants with enhancer activation tags.
[0042] Compared with the wild type, the main feature of slg-D is that the plant type is loose in the vegetative growth period and mature harvest period ( figure 1 A, B), grain length increased, grain width decreased, and thousand-kernel weight decreased ( figure 1 C-F), the leaf angle increases ( figure 1 G, H). The scanning electron microscope observation of the inner and outer epidermal cells of the seed glume shell indicated that the increase in grain length and the decrease in grain width of slg-D grains were caused by the increase in cell length and the decrease in cell width, respectively ( figure 2 A-G); the optical microscope observation of the longitudinal section of the occipital lobe showed that the increase in the in...
Embodiment 2
[0043] In conclusion, the change in cell size in mutant slg-D resulted in slender grains and increased leaf angle phenotypes. Example 2. Cloning and transgenic verification of SLG
[0044] 1. Identification of T-DNA insertion sites
[0045] In order to isolate the mutant gene of the slg-D mutant, the flanking sequence of the mutant number in the T-DNA insertion mutant library was first searched, and three primers were designed: P1, P2, and P3 (Table 1), their positions like image 3 As shown in A, PCR amplification shows that the genome primers P1+P2 can amplify the target band in the wild type, but cannot amplify the band in the slg-D mutant, and the genome vector combination primers P2+P3 in the The target band can be amplified in the slg-D mutant, but the band cannot be amplified in the wild type ( image 3 A, B), proved the authenticity of the T-DNA insertion position, further sequenced the amplified fragment, and passed the data in the Rice Genome Annotation Project Da...
Embodiment 3
[0093] Example 3. Construction of transgenic overexpression vector and Agrobacterium transformation
[0094] Construction of transgenic overexpression vector:
[0095] The pCUbi1390 overexpression vector was digested with KpnI and SpeI double enzymes, electrophoresed, and the linear vector was recovered for future use. The total plant mRNA of the rice variety Dongjin was extracted, and primers were designed according to the CDS sequences predicted on the NCBI website. The 5' ends of the front and rear primers were respectively added with restriction sites and several protective bases. The designed primers 840-1390 F and 840-1390 R to amplify mRNA, electrophoresis, and recover the fragment, then double-enzyme KpnI and SpeI to cut the fragment, electrophoresis, recover the cut fragment, use T4 ligase to connect the fragment to the digested pCUbi1390, and transfer it to the large intestine After the sequence was verified to be correct by sequencing, the plasmid was extracted and...
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