Separated non-expressed promoter SAFES6 in endosperm and application of separated non-expressed promoter SAFES6

A promoter and isolated technology, applied in the fields of biotechnology and crop genetic engineering, can solve problems such as research results having no application value and hindering development

Active Publication Date: 2016-11-23
RICE RES ISTITUTE ANHUI ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although genetically modified crops do have potential food safety and environmental safety problems, they must not be used as a reason to hinder their development, but problems should be discovered and efforts should be made to solve them
If transgenic rice cannot be finally commercialized and used in production, no matter how good the research results are, they will have no application value

Method used

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  • Separated non-expressed promoter SAFES6 in endosperm and application of separated non-expressed promoter SAFES6
  • Separated non-expressed promoter SAFES6 in endosperm and application of separated non-expressed promoter SAFES6
  • Separated non-expressed promoter SAFES6 in endosperm and application of separated non-expressed promoter SAFES6

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] Embodiment 1, the acquisition of SAFES6 promoter

[0064] Step 1. Design of primers

[0065] According to the whole genome sequence of rice variety Nipponbare (Oryza sativa L cv. Nipponbare) provided by NCBI, the amplification primers were designed according to the sequence of the rice promoter SAFES6 (the sequence shown in SEQ ID No: 1 in the sequence table), and according to the selected The characteristics of the vector and the target gene, and the design of the restriction site of the primer. The seeds of the rice variety Nipponbare used in the present invention were obtained from the seed bank of the Rice Research Institute of Anhui Academy of Agricultural Sciences.

[0066] In this embodiment, rice binary expression vector pCAMBIA1391( figure 2 In part A, from CAMBIA, which is a carrier for public use, and is preserved by the rice group of the Supervision, Inspection and Testing Center for GMO Product Components of the Ministry of Agriculture, Anhui Academy of ...

Embodiment 2

[0075] Embodiment 2, the construction of crop expression vector and the transformation of Agrobacterium

[0076] From the Escherichia coli positive clones obtained in the above "acquisition of promoter SAFES6", extract the PGEM-T-Easy vector plasmid containing the promoter SAFES6 fragment, and perform double digestion with HindIII and EcoRI to recover the promoter SAFES6 fragment. At the same time, HindIII and EcoRI were used to digest the crop expression vector pCAMBIA1391, and the pCAMBIA1391 fragment after digestion was recovered. The pCAMBIA1391 vector contains the reporter gene Gus driven by no promoter (the promoter to be studied in the present invention is inserted in the upstream of the Gus gene, for driving the expression of the Gus gene, reflecting the properties of the researched promoter with the expression characteristics of the Gus gene ) The above recovered SAFES6 fragment and pCAMBIA1391 fragment were connected with T4 DNA ligase (purchased from TaKaRa Company)...

Embodiment 3

[0077] Embodiment 3, promoter SAFES6 drives the expression of Gus reporter gene

[0078] Step 1: Agrobacterium-mediated genetic transformation of rice

[0079] After removing the chaff of mature Nipponbare rice seeds, soak the seeds with 70% alcohol for 1 min, and pour off the alcohol. Soak the seeds with a solution of 50% sodium hypochlorite (the concentration of available chlorine in the stock solution is greater than 4%) containing 1 drop of Tween 20 for 40 minutes, and shake them evenly on a shaker (150r / min). Pour off the sodium hypochlorite, wash with sterile water 5 times until the solution is clear, without the smell of sodium hypochlorite. Soak the seeds in sterile water overnight. The embryos were peeled off along the aleurone layer of the seeds with a scalpel, and the embryos were inoculated on callus induction medium. After 11 days of dark culture at 30°C, the callus was separated from the endosperm and germ, and the primary callus in a good state of degerminati...

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Abstract

The invention relates to a separated non-expressed promoter SAFES6 in endosperm and application of the separated non-expressed promoter SAFES6. Particularly, the invention provides the promoter SAFES6 which can transcribe a heterogenous nucleic acid sequence in tissue rather than in the endosperm of a vegetable life, and methods for preparing and using the promoter. The invention also relates to an expression cassette containing the promoter, a recombinant expression vector and a method for obtaining a corresponding transgenic vegetable life. The promoter SAFES6 for paddy rice, which is provided by the invention, can adjust and control genes to be intensively expressed in a non-endosperm part of a plant, but not to be expressed in the endosperm. The separated non-expressed promoter SAFES6 in the endosperm and the application of the separated non-expressed promoter SAFES6 utilize the promoter to modify growth characteristics of the vegetable life in the condition that characters of the endosperm of the vegetable life are not changed, improve the growth performance of the paddy rice, meanwhile, decease edible safety risks of the paddy rice, further, can obtain an ideal transgenic plant, and contribute to the commercial popularization of transgenic paddy rice.

Description

technical field [0001] The invention relates to the technical fields of biotechnology and crop genetic engineering. Specifically, the present invention relates to a rice endosperm non-expression promoter SAFES6 and its application. Background technique [0002] One of the goals of genetic engineering of crops is to develop plants with characteristics or traits desired for agriculture. The technological development of genetic engineering enables researchers to obtain the desired target gene and transform the plant to obtain transgenic plants with desired traits and characteristics. In one aspect, genes can be induced to be expressed in plant cells or plant tissues that do not normally express such genes to confer on these plants a desired phenotype. On the other hand, transcription of a gene or part of a gene in an antisense direction can prevent or inhibit the expression of an endogenous gene to achieve the desired target. Using the promoter to directly control the transc...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/84A01H5/00
CPCC07K14/415C12N15/8205C12N15/8234C12N2800/60C12N15/113
Inventor 魏鹏程杨剑波秦瑞英李娟李莉许蓉芳李浩
Owner RICE RES ISTITUTE ANHUI ACAD OF AGRI SCI
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