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Application of histone methyltransferase sdg723 in regulation of rice heading stage

A technology of heading stage and rice, which is applied in the field of regulating the histone methyltransferase gene of rice heading stage, can solve the problems that the influence of rice growth and development is poorly understood, and achieve the effect of alleviating food shortage

Inactive Publication Date: 2015-10-21
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Currently, little is known about the effects of H3K4 histone methyltransferases on rice growth and development

Method used

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  • Application of histone methyltransferase sdg723 in regulation of rice heading stage
  • Application of histone methyltransferase sdg723 in regulation of rice heading stage
  • Application of histone methyltransferase sdg723 in regulation of rice heading stage

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1: Amplification of full-length cDNA of SDG723 gene

[0035] For the gene SDG723 (gene accession number LOC_Os09g04890) required by the present invention, mainly by RT-PCR method (see: J. Experiment Guide (Third Edition), Beijing, Science Press, 2002 Edition) was amplified to obtain the full-length sequence of the SDG723 gene. The specific operation is as follows:

[0036] 1) Extract RNA from seedling leaves of rice variety Zhonghua 11 (sourced from Institute of Crop Science, Chinese Academy of Agricultural Sciences). The RNA extraction reagent is Invitrogen’s Trizol extraction kit (see the kit’s instruction manual for specific steps) ;

[0037] 2) The steps to synthesize the first strand of cDNA by reverse transcription in RT-PCR are as follows: ① Prepare mixed solution 1: 4 μg of total RNA, DNaseI2U, 10×DNaseI buffer 1 μl, add DEPC (diethyl pyrocarbonate, a strong inhibitor of RNase) Treat water (0.01% DEPC) to 10 μl, mix well and place the mixed solutio...

Embodiment 2

[0043] Example 2: Construction of SDG723 double-strand suppression vector

[0044] The gene cloning of the present invention is carried out by RT-PCR method (see: J. Sambrook, EF Fritsch, T Mani Atis, translated by Huang Peitang, Wang Jiaxi, etc., Molecular Cloning Experiment Guide (Third Edition), Science Publishing House, 2002 edition) to amplify to obtain a specific sequence of SDG723. The specific steps are: according to the full-length cDNA sequence of the SDG723 gene published in the public database (http: / / rice.plantbiology.msu.edu / ), through the ChromDB The database (http: / / www.chromdb.org / index.html) was compared to find the SDG723-specific sequence to design primers and PCR to amplify the RNAi inhibitory fragment. The amplified product was connected to pGEMT-vector (purchased from Promega (Beijing) Biotechnology Co., Ltd., ie Promega, USA) by T / A cloning for sequencing verification.

[0045] The primers used to clone the RNAi suppressor fragment of SDG723 are as fol...

Embodiment 3

[0053] Example 3: Transformation of binary Ti plasmid vector and detection of positive and expression levels of transgenic plants

[0054] 1) The newly constructed suppression expression vector pDS1301-SDG723-2 (from Example 2) was introduced into Agrobacterium EHA105 (purchased From the Australian CAMBIA laboratory) strain, we named the obtained strain pDS1301-SDG723-EHA105.

[0055] 2) Transform the pDS1301-SDG723-EHA105 obtained in the previous step into the japonica rice variety Zhonghua 11. The transformation method refers to the method reported by Hiei et al. (Hiei et al., Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J, 1994, 6:271-282.). The obtained T0 transgenic plants were named Rn, where n=1, 2, 3...represent different transgenic families.

[0056]3) Total DNA was extracted from the leaves of the transformed plants of the T0 generation. The DNA extraction method was the CTA...

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Abstract

The invention discloses application of histone methyltransferase SDG723 in regulation of the heading period of rice, and belongs to the technical field of plant transgene engineering. Experiments show that in transgenic plants restraining the SDG723, H3K4 trimethylation drops significantly, and the fact that the gene is histone H3K4 trimethylation transferase is explained. After the SDG723 is restrained, the heading period of the plants is delayed by about 30 days. The genes related to the heading period are detected by using Realtime-PCR, and the expression quantity of the flowering promoting genes in the plants with the SDG723 restrained is reduced. The mentioned results explain the fact that the SDG723 regulates the heading period of the rice by regulating the H3K4 trimethylation modification.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering. Specifically involving a histone methyltransferase gene that regulates heading date in rice [0002] Isolation cloning, functional verification and application of SDG723. Background technique [0003] Histone modification is one of the important mechanisms of epigenetic regulation. The function of core histone is highly conserved in evolution, but its structure is in dynamic change. Its modification not only affects the expression activity of genes, but also regulates its transcription A change in active or silent state. Methylation of histone lysine is a common histone modification, mainly occurs at lysine 4, 9, 14, 27, 36, 79 of histone H3 and 20 and 59 of lysine of histone H4 (Transcription regulation by histone methylation: interplay between covalent modification of core histone tails. Gene Dev, 2001). In terms of the dynamic changes of the overall chromosome, H3K4 methy...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/54C12N9/10C12N15/84A01H5/00
Inventor 周道绣张华赵毓
Owner HUAZHONG AGRI UNIV
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