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Maize Bract Length Regulatory Protein arr8 and Its Encoding Gene and Application
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一种玉米、蛋白质的技术，应用在植物生物领域，能够解决减产、招虫害、影响灌浆等问题
Active Publication Date: 2021-02-09
CHINA AGRI UNIV
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In recent years, under the influence of high temperature and drought, corn in Liaoning, Heilongjiang, Shanxi, Shandong and other places has appeared the phenomenon of "corn topping". Yield reduction; In addition, short bracts are easy to attract insect damage, such as two-spotted firefly leaf beetle biting filaments and scarabs biting grains, which will also cause yield reduction
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Embodiment 1
[0068] Example 1. Obtaining and sequence analysis of ARR8 protein and its coding gene
[0069] 1. Acquisition of ARR8 protein and its coding gene
[0070] In the study of maize bract phenotype, it was found that GRMZM2G479110 gene plays an important role in regulating the length of maize bract. The GRMZM2G479110 gene (the gene number of the V4 version is Zm00001d018380) is located on chromosome 5, with a total length of 4805 bp, a total of 6 exons, and encoding 676 amino acids. The GRMZM2G479110 gene was named ARR8, the sequence of the ARR8 gene is shown in sequence 1, the CDS sequence of the ARR8 gene is shown in sequence 2, and the amino acid sequence encoded by the ARR8 gene is shown in sequence 3.
[0071] 2. Sequence analysis
[0072] Application of multiple protein domain prediction software (SMART, CDD, PFAM) to search, the results show that: ARR8 gene encoding ARR8 protein contains a highly conserved Myb_DNA_bing domain and a cytokinin response_Reg domain ( figure ...
Embodiment 2
[0073] Example 2, ARR8 gene functional verification
[0074] 1. Obtaining and phenotypic analysis of the deletion mutant of ARR8 gene (crispr-arr)
[0075] 1. Obtaining the deletion mutant (crispr-arr) of the ARR8 gene
[0076] A deletion mutant (crispr-arr) of the maize ARR8 gene was created by CRISPR-Cas9 gene knockout technology combined with the maize gene genetic transformation method. The mutation site is located downstream of the start codon ATG of the first exon protein coding At 28bp, it is a protein coding missense mutation caused by a deletion of 61 bases. Specific steps are as follows:
[0077] 1) Selection of sgRNA sequence
[0078] The target site sequence was designed on the maize ARR8 gene with a length of 20bp.
[0079] The target site 1 is located at positions 1196-1215 of the ARR8 gene sequence (Sequence 1), and the target site 1 sequence is as follows: TGGGGAGAGTCAGGGACCAG.
[0080] The target site 2 is located at positions 1257-1276 of the ARR8 gene s...
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Abstract
The invention discloses the corn bract length regulating protein ARR8 and its coding gene and application. The corn bract length regulatory protein ARR8 is a protein as follows a) or b) or c) or d): a) the amino acid sequence is the protein shown in sequence 2; b) at the N-terminal of the protein shown in sequence 2 and / or a fusion protein obtained by connecting a tag at the C-terminal; c) a protein with the same function obtained by substituting and / or deleting and / or adding the amino acid sequence shown in Sequence 2; d) with The amino acid sequence shown in Sequence 2 has a homology of 75% or more and a protein with the same function. Through the verification of transgene knockout and ARR8 mutants, the length of bracts in ARR8 gene deletion mutants and ARR8 mutants was significantly increased, indicating that ARR8 plays an important role in regulating the length of bracts.
Description
technical field [0001] The invention relates to the field of plant biotechnology, in particular to the maize bract length regulating protein ARR8 and its coding gene and application. Background technique [0002] Machine harvesting of corn is a major change in the planting mode, and it is a key measure and development trend to achieve high-yield and high-efficiency production of corn in large areas. However, the corn varieties currently popularized in my country generally have the problems of too slow grain dehydration and high water content during harvest, which seriously restricts the industrialization process of corn combined harvesting. Factors affecting the water content of maize at maturity include variety maturity, ear bract traits, and grain dehydration rate at the late stage of grain filling; among them, bract traits directly affect ear dehydration, and indirectly affect grain water loss, which is the key factor for high-yield and suitable corn grains. Key traits t...
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