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ZmAGO1a protein as well as coding gene and application thereof

A technology that encodes genes and proteins, which can be used in applications, genetic engineering, plant genetic improvement, etc., and can solve problems such as lack of evidence at the molecular level.

Inactive Publication Date: 2015-05-27
INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Qian Yexiong et al. used bioinformatics methods to predict 18 ZmAGO genes, including five members of ZmAGO1, but there is still a lack of evidence at the molecular level

Method used

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  • ZmAGO1a protein as well as coding gene and application thereof
  • ZmAGO1a protein as well as coding gene and application thereof
  • ZmAGO1a protein as well as coding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1, Preparation of Plant Overexpression Vector pCPB-35S::ZmAGO1a

[0049] 1. The full-length cDNA sequence of the ZmAGO1a gene of the maize inbred line Ye 478 is shown in SEQ ID No. 1, and the 150th to 3458th positions in SEQ ID No. 1 are the coding gene sequence of the ZmAGO1a protein. The amino acid sequence of the ZmAGO1a protein is shown in SEQ ID No.2.

[0050] 2. Construction of pCPB-35S::ZmAGO1a

[0051] The sequence shown in the 123rd to the 3610th nucleotide in SEQ ID No.1 was inserted into the BamHI site of pCPB, and the remaining sequences of pCPB remained unchanged to obtain a recombinant plasmid, which was named pCPB-35S::ZmAGO1a, The pCPB-35S::ZmAGO1a was sent for sequencing, and the result was correct. The sequencing results showed that the sequence shown in the 123rd to 3610th nucleotides in SEQ ID No.1 was correctly inserted into the BamHI site of pCPB, and SEQ ID No. 2 shows the ZmAGO1a protein.

[0052] The schematic diagram of the plant ove...

Embodiment 2

[0053] Example 2. Functional complementation identification of Arabidopsis ago1 mutant by ZmAGO1a gene

[0054] 1. After sterilizing the seeds of the Colombian ecotype Arabidopsis thaliana and the Arabidopsis ago1-27 mutant, they were placed in 1 / 2MS (Murashige and Skoog) medium respectively, under short-day light (8h light / 16h dark), 23 Cultivate at ±1°C for 10 days, then transfer to soil, and cultivate under short-day light (8h light / 16h dark) at 23±1°C.

[0055] 2. Transform the plant overexpression vector pCPB-35S::ZmAGO1a prepared in Example 1 into Agrobacterium GV3301 to obtain recombinant Agrobacterium, and use the inflorescence infiltration method (Clough SJ, et al., 1998) to infect the recombinant Agrobacterium A cultured Colombian ecotype Arabidopsis thaliana and Arabidopsis ago1-27 mutant, respectively obtained Columbia ecotype Arabidopsis transgenic plants and Arabidopsis ago1-27 mutant transgenic plants, and harvested Columbia ecotype Arabidopsis transgenic plants...

Embodiment 3

[0099] Example 3, Tissue-specific analysis of ZmAGO1a gene expression in different organs of maize

[0100] 1. Collect the tassels at the tasseling stage, silk at the silking stage, bracts at the silking stage, and ear ears at the silking stage of the corn inbred line Ye 478 that grows normally in the field, and three of the normal hydroponic corn inbred line Ye 478. Leaf sheaths, young roots, and young leaves at the leaf stage.

[0101] 2. Extract the total RNA of each tissue collected in step 1, and reverse transcribe to obtain cDNA, respectively use the extracted cDNA as a template, and use ZmAGO1a forward primer (SEQ ID No.7) and ZmAGO1a reverse primer (SEQ ID No. .8) Perform real-time fluorescent quantitative PCR as primers to detect the relative expression level of the ZmAGO1a gene, or use the GAPDH forward primer and GAPDH reverse primer as primers to perform real-time fluorescent quantitative PCR to detect the relative expression level of the internal reference gene GA...

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Abstract

The invention discloses ZmAGO1a protein as well as a coding gene and application thereof. The protein disclosed by the invention is represented as follows: (1), protein represented by SEQ ID No.2; and (2), protein obtained by substitution and / or deletion and / or addition of an amino acid sequence represented by SEQ ID No.2 through one or more amino acid residues and having same functions. The ZmAGO1a gene disclosed by the invention is capable of obviously influencing growth and development of plants, such as fecundity, a maturation period (early maturation), florescence, the number of leaves, the sizes of the leaves, the shapes of the leaves and the colors of the leaves; most of the characteristics belong to important agronomic characteristics and are related to the yield, growth period and plant type of plants; and the ZmAGO1a protein disclosed by the invention has important application value in cultivation of high-yield, stress-tolerant and high-efficiency plants (such as corns).

Description

technical field [0001] The invention relates to a ZmAGO1a protein and its coding gene and application, belonging to the field of biotechnology. Background technique [0002] As a core element of the RNA-induced silencing complex, AGO protein plays a key role in RNA silencing. Prokaryotic AGO protein is a key component in the defense system against the invasion of foreign genetic elements, and archaea has similar functions in the RNAi system. Eukaryotic AGO proteins play an important role in the regulation of gene expression and are widely involved in a variety of biological processes, including developmental timing, cell differentiation, cell proliferation, cell death, metabolic control, immunity, and transposon silencing (Wei KF, et al ., 2012, Pumplin N, et al., 2013), in addition, AGO protein plays an important role in transcription, alternative splicing, and even DNA repair (Meister G, 2013). AGO is a multi-domain, larger protein (approximately 90-100 kDa). Eukaryotic...

Claims

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Application Information

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IPC IPC(8): C07K14/415C12N15/29C12N15/82A01H5/00C12N1/15C12N1/21C12N1/19
CPCC07K14/415C12N15/8205C12N15/8261
Inventor 谢传晓徐东东李新海李猛王述民刘方王慧
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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