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Histone demethylase gene JMJ703 for regulating rice stem height, and applications thereof

A rice and protein technology, applied in the field of plant genetic engineering, can solve the problems of less research

Inactive Publication Date: 2014-07-16
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Jumonji-like proteins are the main family of the only two histone demethylase families discovered so far, and they are widely involved in the regulation of gene expression, the formation of chromatin structure and the development of organisms. There are very few studies in

Method used

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  • Histone demethylase gene JMJ703 for regulating rice stem height, and applications thereof
  • Histone demethylase gene JMJ703 for regulating rice stem height, and applications thereof
  • Histone demethylase gene JMJ703 for regulating rice stem height, and applications thereof

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Embodiment 1

[0040] Example 1: Obtaining related sequences

[0041] For the gene required for the present invention (accession number AK121381, http: / / www.ncbi.nlm.nih.gov / nucleotide / 37991004), mainly by RT-PCR method (see: J. Sambrook, EF Fritsch, T Mani Artis, translated by Huang Peitang, Wang Jiaxi, etc., Molecular Cloning Experiment Guide (Third Edition), Beijing, Science Press, 2002 edition) was amplified to obtain a specific sequence of JMJ703 gene,

[0042] A. First extract the RNA from the leaves of rice seedlings. The reagent for RNA extraction is Trizol extraction kit from Invitrogen Company (see the kit manual for specific operation steps);

[0043] B. The steps to synthesize the first strand of cDNA by reverse transcription in RT-PCR are as follows: ① Prepare mixed solution 1: 2 μg of total RNA, 2U of DNaseI, 1 μl of 10x DNAseI buffer, add DEPC (diethyl pyrocarbonate, strong inhibitor of RNase solution) treated water (0.01% DEPC) to 10 μl, after mixing, place the mixed solutio...

Embodiment 2

[0053] Embodiment 2: Construction of RNAi double-strand suppression expression vector:

[0054] 1) Digest the obtained sequence of SEQ ID NO: 1 with BamHI and KpnI (purchased from Bao Bioengineering Dalian Co., Ltd., see the specification of the endonuclease for details), recover the target fragment and connect it to the double-strand suppression vector. The ligase was purchased from NEB Company, and the reaction system was referred to the instructions.

[0055] 2) The ligation product was introduced into Escherichia coli DH10B (purchased from Promega (Beijing) Biotechnology Co., Ltd., the United States) by electrotransformation (the electrotransformer is a product of eppendorf, and the voltage used is 1800v, ​​see the instrument manual for the operation method). Promega Company), in LA containing 250ppm kanamycin (purchased from Roche Biological Company) (LA formula see J. Molecular Cloning Experiment Guide (Third Edition), Science Press, 2002 Edition) cultured on LA-resista...

Embodiment 3

[0063] Example 3: Function of a rice histone demethylase in histone H3K4 demethylation

[0064] A. Analysis of demethylase activity of JMJ703 protein in vivo:

[0065] Since histone methylation modification can occur in different forms in different sites, and the demethylation activity of JMJ proteins is also specific, so the demethylation activity of JMJ703 protein was determined by heterologous expression in tobacco. The nucleotide sequence shown in the sequence table as SEQ ID NO: 2 was cloned into the tobacco expression vector pFA121 (see Figure 1), and the expression of JMJ protein was detected by Immunostaining after transforming tobacco with Agrobacterium histone modifications in the nucleus. The specific method is as follows:

[0066] 1) Use XbaI to single-digest the nucleotide sequence shown in SEQ ID NO: 2 in the list, recover the target fragment, and clone it into pFA121 (see Figure 1). After the 35S primer is sequenced to verify the correct direction, use The me...

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Abstract

The invention discloses a histone demethylase gene JMJ703 for regulating a rice stem height, and applications thereof. According to the present invention, one of histone demethylases and gene JMJ703 thereof are selected through searching in a chromatin protein database, wherein the full-length cDNA accession number is AK121381, a gene ID number is 4338043, a prediction coding region comprises 3717 bp, and 1238 amino acids are coded; a RNA interference vector of the gene is transformed into rice through a RNA interference method, and results show that a stem height of the transgenic plant is shorter than a stem height of the wild type while a length of the leaf has no apparent change so as to improve a column shape of the rice; tobacco heterogenesis in vivo test results show that JMJ703 protein is demethylase of histone H3K4me1 / H3K4me2 / H3K4me3; and the JMJ703 is functional histone demethylase gene and can be provided for regulating a stem elongation process of the rice, wherein the stem of the rice expressed after gene inhibition becomes short and a characteristic of lodging resistance is provided while developments of leaves and other organs are normal so as to provide a selective plant type for breeding.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering. It specifically relates to a histone demethylase gene JMJ703 that regulates the stem height of rice, and also relates to a preparation method for the histone demethylase gene JMJ703 that regulates the stem height of rice, and also relates to a method for regulating rice stem height Use of the stalk height histone demethylase gene JMJ703, which is involved in the regulation of histone methylation and plant stalk height. Background technique [0002] Food security is already a worldwide problem. As one of the main food crops in the world, rice is of great significance to alleviating food shortages, especially ensuring food security in my country. How to increase rice production has become a scientific issue of great significance. At the same time, as a model organism for Poaceae research, related research also has considerable theoretical significance. In the early 1960s, the r...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/53C12N9/02C12N15/84A01H5/00
Inventor 周道绣陈香嵩
Owner HUAZHONG AGRI UNIV
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