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Transcription factor gene OsHOX4 for controlling synthesis of rice gibberellin and use thereof

A technology of transcription factor and gibberellin, which is applied in the genetic engineering of rice and the field of new plants, can solve the problems that the function needs to be further studied.

Inactive Publication Date: 2008-12-24
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the classification of plant species, Arabidopsis is a dicotyledonous plant and maize is a monocotyledonous plant. Therefore, whether this gene can transform maize and regulate the synthesis of gibberellin in maize plants remains to be further studied.

Method used

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  • Transcription factor gene OsHOX4 for controlling synthesis of rice gibberellin and use thereof
  • Transcription factor gene OsHOX4 for controlling synthesis of rice gibberellin and use thereof
  • Transcription factor gene OsHOX4 for controlling synthesis of rice gibberellin and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Cloning of candidate gene OsHOX4:

[0033] Using Arabidopsis AtHB6 (Homeobox (HOX) class transcription factor, NCBI protein accession number AAD41726; in the REDB website ( http: / / redb.ncpgr.cn / ), select the Regularblast tool in the "Rice EST" database for tblastx analysis, and find the homologous clone in rice (numbered as EI#77-F09), from the cDNA library (see references: Chu Zhaohui, Peng Kaiman, etc., the whole growth period of rice Construction and identification of a homogeneous cDNA library, Science Bulletin, 2002, 47(21).1656-1662) obtained this clone, sequenced to obtain a full-length cDNA sequence (sequence determination was completed by the Shanghai National Gene Sequencing Center), and named the candidate gene It is OsHOX4, and its cDNA nucleotide sequence is shown in SEQ NO: 1 in the sequence table. Use GenDoc software (version is selected from: GenDoc3.2) to the protein sequence analysis of the gene OsHOX4 of reported AtHB6 and the present inv...

Embodiment 2

[0034] Example 2 The construction of the binary Ti plasmid vector and the establishment of transformed Agrobacterium:

[0035] Specific steps are as follows:

[0036] 1) Cloning the cDNA with OsHOX4, loading the plasmid as pSPORT1 (purchased from Promega, a special plasmid vector for constructing cDNA library) digested with KpnI and BamHI, and then isolating the target gene fragment (plus the enzyme derived from pSPORT1 Cut the linker fragment, the size of the cloned OsHOX4 gene is 1560bp), and directly connect with the vector plasmid pDS1301 digested with KpnI and BamHI (the endonucleases used are all purchased from TAKARA company, and the specific usage and dosage refer to the product manual; The ligase was purchased from Invitrogen, and the usage and dosage refer to the instructions of the product);

[0037] 2) The connection product is introduced into DH10B (purchased from Promega Company) by electrotransformation (the electrotransformer is a product of eppendorf, and the...

Embodiment 3

[0040] Example 3 Transformation of binary Ti plasmid vector and positive detection of transgenic plants:

[0041] Specific steps are as follows:

[0042] 1) The transformed strain T-HOX4 obtained in Example 2 was introduced into the recipient rice variety "Nipponbare", and the transformation method was referred to the method reported by Hiei et al. (see: Efficient transformation of rice, Oryza sativa L., mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA, 1994, Plant Journal 6:271-282). The obtained T0 transgenic rice plants were named HOX4-n, wherein n=1, 2, 3...represent different transgenic families.

[0043] Total DNA was extracted from leaves of transformed rice plants of the T0 generation. The DNA extraction method is the CTAB method (with reference to Zhang et al., the method and steps shown in genetic diversity and differentiation of indica an japonica rice detected by RFLP analysis, 1992, Theor Appl Genet, 83, 495-499), and then PCR metho...

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Abstract

The invention belongs to the technical field of plant genetic engineering. It specifically relates to the isolation and cloning, functional verification and application of a DNA fragment (gene) regulating rice gibberellin synthesis. The gene OsHOX4 is related to the synthesis of gibberellin in rice. The complete translation sequence (Coding sequence) of the gene was combined with the cauliflower mosaic virus promoter (CaMV35S) and directly transformed into rice, and the transgenic plants were significantly shorter than the wild-type control plants; under the action of exogenous gibberellin (GA3), the transgenic The plant height returned to normal, and further analysis found that the expression of gibberellin oxidase gene (OsGA3ox2) in the transgenic plants was inhibited. It shows that OsHOX4 can regulate the expression of OsGA3ox2 and then control the synthesis of gibberellin.

Description

technical field [0001] The invention belongs to the technical field of new plants, in particular to the technical field of genetic engineering of rice. A transcription factor gene OsHOX4 that regulates gibberellin synthesis in rice was isolated, the function of the gene was verified and its application in transgenic rice. Background technique [0002] Gibberellin (GA) belongs to diterpenoids, and more than 70 species have been isolated from higher plants and microorganisms. All gibberellins have a common basic skeleton in chemical structure, that is, gibberellane, and all contain carboxyl groups, so they are acidic. The structural differences between various gibberellins mainly lie in the number and position of double bonds and hydroxyl groups. [0003] In higher plants, almost all tissues and organs contain gibberellin, but the content is generally very small, and the content is often higher in vigorously growing parts. The main parts of gibberellin synthesis in plants a...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82
Inventor 代明球周道绣张启发
Owner HUAZHONG AGRI UNIV
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