Promoter OsP002, preparation method and applications thereof

A promoter and plant technology, applied in the field of plant genetic engineering, can solve problems such as limited regulatory effect

Inactive Publication Date: 2014-03-05
华大基因杨凌创新研究院有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The Adh-1 promoter is mainly used in monocotyledonous plants, and has limited effect on the regulation of gene expression in most dicotyledonous plants

Method used

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  • Promoter OsP002, preparation method and applications thereof
  • Promoter OsP002, preparation method and applications thereof
  • Promoter OsP002, preparation method and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] Example 1: PCR amplification of P002 promoter fragment and construction of pMD18-T+P002 recombinant vector.

[0066] 1. PCR amplification of the P002 promoter fragment:

[0067] Using the Plant Genomic DNA Extraction Kit (TIANGEN New Plant Genomic DNA Extraction Kit, catalog number: DP320-02) to extract rice Nipponbare (the application number is

[0068]200910238992.0, the invention title is "A kind of promoter BgIosP587, its preparation method and application" is preserved in the invention application, and published on September 22, 2010, the preservation number is CCTCC NO: P200910) Genomic DNA, according to the promoter In the sequence in rice Nipponbare gDNA, design a pair of PCR specific amplification primers at the beginning and end respectively (upstream primer F1, plus restriction enzyme cutting site EcoR I and protection base, downstream primer R1, plus restriction enzyme cutting site SbfI and protected bases). Using the gDNA of rice Nipponbare extracted abov...

Embodiment 2

[0079] Example 2: Construction of vector-p8+P002 recombinant vector.

[0080] 1. Construction of p8 plasmid:

[0081] The p8 plasmid used in the present invention is provided by pCAMBIA-1301 plasmid (Dong Yang, Kunming Institute of Zoology, Chinese Academy of Sciences; or can be purchased from, for example, Shanghai Guorui Gene Technology Co., Ltd., the original source of the company's pCAMBIA-1301 plasmid is The CAMBIA Bios (biological open source) Licensee, Australia) was modified and constructed in the following way, and the specific instructions are as follows:

[0082] use as figure 1 The indicated Kpn I / Nco I (NEB) double enzyme digested plasmid pCAMBIA-1301, and recovered large fragments. Synthesize the following sequence according to the restriction endonuclease sites employed:

[0083] GGTACCAAGCTTACTAGTCCTGCAGGTCTAGAGGATCCGTCGACCATGG (SEQ ID NO: 6) (restriction site included is Kpn I / HindIII / Spe I / Sbf I / Pst I / Xba I / BamH I / SalI / Nco I), with Kpn I / Nco I double enzym...

Embodiment 3

[0101] Example 3: Preparation of recombinant Agrobacterium tumefaciens EHA105-P002 cells.

[0102] The p8+P002 recombinant vector constructed as described in Example 2 and the p8 plasmid as a control were respectively transformed into root cancer prepared by the calcium chloride method described in the "Molecular Cloning Experiment Guide" (third edition, Science Press). Agrobacterium EHA105 (has been preserved in the invention application with the application number 200910238992.0 and the invention name "A promoter BgIosP587, its preparation method and use", and was published on September 22, 2010, and the preservation number is CCTCC NO: M209315 ) competent cells, the specific method is as follows: take out the competent cells of Agrobacterium tumefaciens EHA105 from the ultra-low temperature refrigerator, and thaw on ice. After thawing, add 5 μl of p8+P002 recombinant vector and p8 plasmid and p8 empty vector as a control, mix gently, ice bath for 10 minutes, freeze in liqui...

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Abstract

The invention provides a promoter OsP002, a preparation method and applications thereof. The nucleotide sequence of the promoter is represented by the SEQ ID No.1, or the nucleotide sequence of the promoter is in a complementation relationship with a nucleotide sequence represented by the SEQ ID No.1. The promoter can be used to modulate the expression of the target gene in monocotyledons, and provides a novel tool and choice for researching the expression of target gene in monocotyledons.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, and specifically relates to a promoter OsP002, a preparation method and an application thereof. Background technique [0002] Promoter refers to a DNA sequence in a gene that can combine with RNA polymerase and other trans-factors that affect transcription to achieve precise and effective initiation of transcription. The start time and degree of plant gene expression (transcription) are controlled by the promoter. The cloning and functional analysis of the promoter is an important part of the study of plant gene expression regulation, and it is also an important aspect of plant genetic engineering research. [0003] Promoters can be divided into three categories according to their transcriptional patterns: constitutive promoters, tissue or organ-specific promoters, and inducible promoters. The so-called constitutive promoter means that under the regulation of the constitutive p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/70C12N15/84C12N1/21C12N5/10A01H5/00C12N15/11C12N15/10
Inventor 黄刚汪杰焦伟钢张厚宝辛宜拓庆荣田伯瑞廖加涛杨爽
Owner 华大基因杨凌创新研究院有限公司
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