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Paddy istone lysine methyltransferase, coding genes thereof and application thereof

A technology of lysine methylation and histone, applied in the direction of transferase, application, genetic engineering, etc., to achieve high application value and broad application prospects

Inactive Publication Date: 2013-01-02
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the long life cycle of rice and other reasons, the relevant research is still quite lagging behind compared with Arabidopsis, and the research on the methylation modification of histone H3K36 in rice is still blank.

Method used

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  • Paddy istone lysine methyltransferase, coding genes thereof and application thereof
  • Paddy istone lysine methyltransferase, coding genes thereof and application thereof
  • Paddy istone lysine methyltransferase, coding genes thereof and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1. Acquisition of rice histone lysine methyltransferase gene SDG725

[0031] The SDG8 gene sequence of Arabidopsis thaliana published in GenBank (GenBank number: NM_106379) was compared with the homologous sequence, and the homologous sequence was obtained from the rice genome, and a sequence was designed based on the 5' and 3' end sequences of the homologous sequence. For the primers, the primer sequences were: 5'-ATGGAGGAGCCTGACGGGGA-3', and 5'-TCAAAATCTTTGATTATTGTTAGGA-3'.

[0032] Extract the total RNA of rice yellow seedlings (Promega, SV total RNA isolation system), use the total RNA of rice as a template, and use AMV reverse transcriptase (TaKaRa) to synthesize cDNA (according to the user manual of Plant RT-PCR Kit 2.01 (TaKaRa) conduct). Using cDNA as a template, the full-length cDNA sequence of rice histone lysine methyltransferase gene SDG725 was amplified by PCR. The 50ul PCR reaction system contains: template 2ul, high-fidelity enzyme KOD plus (TOY...

Embodiment 2

[0033] Example 2. Determination of protein activity of rice histone lysine methyltransferase gene SDG725

[0034]Using the pUC19-SDG725 obtained in Example 1 as a template, PCR amplifies the nucleotide sequence at position 3646-4656 from the 5' end of SDG725 (including the active center of histone lysine methyltransferase—the SET domain), The 5' and 3' primers were: 5'-acggatccATGAATAAACAGAGTGATCTTATTC-3', and 5'-actcgagTCAACCACCGATGTAACCCCGG-3', respectively. The 50 μl PCR amplification system contained template pUC19-SDG725200ng, 20 pmol of 5' and 3' primers, 5 μl of 10X buffer, 20 μM of dNTP, and 1ul of high-fidelity KOD plus (TOYOBO). Amplification was performed under the following conditions: pre-denaturation at 94°C for 2 minutes; denaturation at 94°C for 30 seconds, annealing at 55°C for 30 seconds, and extension at 68°C for 1 minute, a total of 30 cycles. The PCR product was cloned into the Escherichia coli expression vector pGEX-4T1 (GE life sciences) through BamH I ...

Embodiment 3

[0038] Example 3. Detection of the regulatory effect of SDG725 on rice growth and development

[0039] 1. Construction of SDG725 antisense expression vector

[0040] Using RNA interference (RNA interference, RNAi) technology, the SDG725 gene was used as the target gene to construct the RNA interference vector. Using the pUC19-SDG725 obtained in Example 1 as a template, two pairs of different primers were used to PCR amplify the nucleotide sequence at positions 3639-3745 from the 5' end of SDG725 to form a complementary DNA double strand with a hairpin structure. The primer pair 1 used was: 5'-aaggatccACCTCGCATGAATAAACAGAG-3' and 5'-aaaagcttCAGTTTTGCAAGCAACAACTG-3', and the obtained PCR product was fSDG725i (forward). Primer pair 2 is 5'-aagaattcCAGTTTTGCAAGCAACAACTG-3' and 5'-aaactagtACCTCGCATGAATAAACAGAG-3', and the resulting PCR product is rSDG725i (reversed). In order to facilitate the subsequent vector construction, restriction enzyme sites of BamH I and HindIII were add...

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Abstract

The invention belongs to the technical field of biological histone genes, and particularly discloses paddy histone lysine methyltransferase, coding genes thereof and application thereof. The methyltransferase has one of amino acid residue sequences, namely 1) an amino acid residue sequence SEQ ID No:1 in a sequence table; and 2) an amino acid residue sequence formed by substituting and / or losing and / or adding 1 to 50 amino acid residues in the amino acid residue sequence SEQ ID No:1 in the sequence table. The methyltransferase is a protein that has the regulation and control effect on the growing development of plants. Antisense transgenic strains of the coding genes of the histone lysine methyltransferase of the paddy are short and small in growth. The histone lysine methyltransferase provides the high-quality genes for the improvement of plant varieties, and has high practical application value and wide application prospects.

Description

technical field [0001] The invention belongs to the technical field of biological histone genes, and in particular relates to a histone lysine methyltransferase related to rice epigenetic regulation and its coding gene and application, in particular to the role of the protein and its gene in regulating plant growth developmental applications. Background technique [0002] The processes of plant growth and development and stress response are all related to the regulation of gene transcription and expression. The regulation of gene transcription and expression is not only affected by genetic factors but also regulated by epigenetics. In recent years, epigenetics has become a hot field of life science research. In addition to the DNA sequence in the genome, there is also a lot of information that regulates genes. They do not change the sequence of the gene itself, but can affect and regulate the function and characteristics of the gene through genetic modification, and inher...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/10C12N15/54C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21C12N15/11A01H5/00
Inventor 沈文辉朱炎叶盛高娟隋鹏飞金菁俞瑜董爱武
Owner FUDAN UNIV
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