Plant endosperm specific expression promoter and use thereof

A specific, promoter technology, applied in applications, plant products, botanical equipment and methods, etc., can solve the problems of consuming bioenergy, the expression intensity cannot meet the needs of transgenic industrialization, and the growth and development of organisms adversely affect metabolites, etc. , to achieve great application prospects, increase the added value of science and technology, and improve the quality of seeds

Inactive Publication Date: 2011-11-23
INST OF BOTANY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although the widely used promoters (including CaMV35S, ubiquitin1, and Actin1) have high expression efficiency, they can express foreign genes in almost all tissues because their expression is not limited by time and space. During the expression process, it will drive the expression of genes in other tissues outside the seed, which will not only consume bioenergy, but also lead to the synthesis of some metabolites that may adversely affect the growth and development of organisms
Moreover, the expression intensity of the above-mentioned promoters cannot meet the needs of transgenic industrialization.

Method used

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  • Plant endosperm specific expression promoter and use thereof
  • Plant endosperm specific expression promoter and use thereof
  • Plant endosperm specific expression promoter and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Embodiment 1, the acquisition of plant endosperm-specific expression promoter (GluB-6 promoter)

[0038] According to the report of Qu et al. (J. Exp. Bot. 2008, 59: 2417-2424), a glutenin gene (gene name Os02g0248800) was selected and named GluB-6. Search the upstream sequence of GluB-6 gene from GenBank, and design primers to amplify the GluB-6 promoter. To facilitate vector construction, two restriction sites (underlined) were sequentially added to the primers. The forward primer of the promoter is GluB-6HindF: 5'-TT AAGCTT TTCGCTGGATTAGAATTC-3'( Hind III ), the reverse primer is GluB-6SalR: 5'GC GTC GAC AGCTTTTGTATATACTAATAAAAC-3'( Sal I ).

[0039] The CTAB method was used to extract a small amount of genome from the leaves of rice (wild type rice Taichung No. 65, which was bred in Taiwan Province of my country in 1936; Qu Leqing et al., Acta Botany, 2001, 43: 1167-1171; preserved by the Institute of Botany, Chinese Academy of Sciences) Using the DNA as a ...

Embodiment 2

[0040] Embodiment 2, plant endosperm-specific expression promoter (GluB-6 promoter) expression vector construction and genetic transformation

[0041] 1. Construction of plant expression vector with GluB-6 promoter fused to GUS gene

[0042] The binary expression vector pGPTV-35S-HPT was constructed according to the method described in the literature (Qu and Takaiwa, Plant Biotech J 2004, 2: 113-125), and was preserved by the Institute of Botany, Chinese Academy of Sciences. The pMD-GluB-6 plasmid and pGPTV-35S-HPT were digested with Sal I and Hind III. A 2192 bp restriction fragment containing the GluB-6 promoter was recovered, and this fragment was ligated between the Sal I and Hind III enzyme recognition sites of pGPTV-35S-HPT. On the basis of PCR identification, the obtained recombinant plasmid was identified by double enzyme digestion with Sal I and Hind III, and a fragment containing 2.2 kb GluB-6 promoter was obtained. The double-enzyme digestion identification shows ...

Embodiment 3

[0047] Example 3, Functional verification of plant endosperm-specific expression promoter (GluB-6 promoter)

[0048] Transgenic GluB-6-pGPTV-35S-HPT rice T 0 Plants were subjected to histochemical staining. Specific steps: Transplant GluB-6-pGPTV-35S-HPT rice T 0 Part of the leaves, roots, stem tissues of the generation plants, and the seeds at the filling stage 11 days, 15 days, and 17 days after flowering that were cut longitudinally from the middle with a scalpel were soaked in GUS reaction solution (0.1M NaPO 4 Buffer, pH7.0, 10mM EDTA, pH7.0, 5mM potassium ferricyanide, 5mM potassium ferrocyanide, 1.0mMX-Gluc, 0.1% Triton X-100), react at 37°C. The stained tissues were preserved in 70% ethanol, observed, and photographed under a dissecting microscope. The results showed that GUS expression was not observed in the roots, stems and leaves of transgenic GluB-6-pGPTV-35S-HPT rice. Image 6 Shown; the outer endosperm of the 11-day-after-anthesis seeds turned blue, but the ...

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Abstract

The invention discloses a plant endosperm specific expression promoter and a use thereof. A nucleotide sequence of the promoter is one of the following nucleotide sequences of 1) a DNA sequence limited by sequence 1 shown in a sequence list in the patent specification, 2) a DNA sequence which is above 70% homologous to the DNA sequence limited by the sequence 1 shown in the sequence list and has functions the same as that of the DNA sequence limited by the sequence 1 shown in the sequence list, and 3) a nucleotide sequence which can hybridize with the DNA sequence limited by the sequence 1 shown in the sequence list under highly stringent conditions. The promoter can promote specific expression of a foreign gene in plant endosperm, thus is suitable for any plant of which a seed has endosperm. In other words, the promoter is suitable for monocotyledons or endosperm type dicotyledons.

Description

technical field [0001] The invention relates to a plant endosperm specific expression promoter and application thereof. Background technique [0002] The latest development of plant biotechnology not only realizes the improvement of traditional agronomic shapes (such as increasing crop yield, enhancing disease resistance, insect resistance, herbicide resistance, improving quality, etc.), but also makes plants become bioreactors for biomedicine and industrial products (Daniell et al., Trends Plant Sci. 2001, 6: 219-226; Giddings et al., Nature Biotech. 2000, 18: 1151-1156; Hood and Jilka, Curr. Opin. Biotechnol. 1999, 10: 382-386 ; Hood and Woodard, Plants as Factories for Protein Production 2002, pp.119-135. Netherlands: Kluwer Academic). For most gramineous crops, due to its high yield, low production cost, storage resistance and easy control of production scale, it is an ideal carrier for the second generation of transgenic products. In recent years, the use of rice seed...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21A01H5/00
Inventor 曲乐庆徐秀萍董祥柏柴志坚
Owner INST OF BOTANY CHINESE ACAD OF SCI
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