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Rice root-specific promoter Os03g01700 and application thereof

A specific promoter, rice root technology, applied in the field of genetic engineering

Inactive Publication Date: 2013-02-27
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] There are no related reports about the rice Os03g01700 promoter regulating the root-specific expression and application of exogenous genes

Method used

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  • Rice root-specific promoter Os03g01700 and application thereof
  • Rice root-specific promoter Os03g01700 and application thereof
  • Rice root-specific promoter Os03g01700 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Embodiment 1, the screening of root specific expression gene:

[0069] 1. Sample preparation

[0070] Pick plump Nipponbare wild-type rice seeds, shell them, wash them with clean water, and then soak them in clean water. Soak the seeds at 37°C until they are white (1-2 days), and change the water in the morning and evening. The germinated seeds are directly broadcast in 3L rice full nutrient solution (pH value 5.5), the formula of the rice full nutrient solution mother solution is shown in (Table 1), the specific preparation of the rice full nutrient solution is as follows, that is, add I-VI number 5ml each stock solution, adjust the pH value to 5.5. It was grown for 10 days in a rice light-temperature-controlled growth room (30° C. during the day, 22° C. at night, light intensity 3000 lux, light duration 12 hours). The shoots and roots were sampled separately. Another spikelet of rice grown in normal nutrient solution to the heading and flowering stage was taken and...

Embodiment 2

[0132] Example 2, expression analysis of rice root-specific promoter Os03g01700 regulating GUS reporter gene

[0133]1. Construction of rice root-specific promoter fusion GUS vector

[0134] The Os03g01700 promoter primer was designed according to the rice genome sequence (TIGRE), and the restriction sites HindIII (shown in lowercase font), BamHI (shown in lowercase italic font) and protective bases (shown in underline "_") were added.

[0135] Os03g01700-3.5Kb promoter primer

[0136] F: 5'- aagcttTGAAAAGAGCTTGAGAGAAT-3'

[0137] R: 5'- ggatccCTTCTTTCTTCTTCGATCGAC-3'

[0138] Using genomic DNA as a template, the OsO3g01700 promoter was amplified with phusion ultra-fidelity DNA polymerase, with a full length of 3500 bp, as shown in SEQ ID NO:1. After recovery by electrophoresis, the PCR product recovered by double digestion with HindIII and BamHI was connected into the GUS plus vector ( Figure 6 , Figure 7 ), to get Os03g01700 promoter: GUSplus. Transform DH5α comp...

Embodiment 3

[0151] Example 3, the Os03g01700 promoter regulates the specific expression of OsPT2 in rice roots

[0152] 1. Os03g01700 promoter regulates the construction of OsPT2 vector

[0153] Use primers to construct the Os03g01700-3.5Kb promoter::GUS vector, introduce EcoR I (shown in lowercase fonts), SmaI (shown in lowercase italic fonts) restriction sites, and add protection bases (shown in underlined "_" fonts) design Os03g01700-3.5Kb promoter double restriction primer; according to the GAL4 / VP16+UAS sequence on the pFX-E24.2R plasmid, introduce the Sma I restriction site to design the GAL4 / VP16+UAS In-Fusion primer ( http: / / bioinfo.clontech.com / infusion / ); according to the OsPT2 (AF536962) information published on NCBI, determine the OsPT2 genome sequence (it has no intron), design In-Fusion primers, and introduce BamHI. The primer sequences are as follows:

[0154] Os03g01700-3.5Kb promote (EcoR I, SmaI)

[0155] F: 5' gaattcTGAAAAGAGCTTGAGAGAAT-3'

[0156] R: 5'- TCC ...

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Abstract

The invention discloses a rice root-specific promoter Os03g01700 and application thereof. The rice root-specific promoter Os03g01700 has the nucleotide sequence shown as SEQ ID NO:1. The rice root-specific promoter Os03g01700 can be used for constructing transgenic plant vectors with root-specific expression. The invention also provides a transgenic plant cell, which comprises the promoter shown as the SEQ ID NO:1. By regulating and controlling the specific expression of exogenous gene in a root through the promoter, the drought resistance and salt and alkali tolerance of the plants can be improved.

Description

technical field [0001] The present invention belongs to the field of genetic engineering, specifically, the present invention relates to the promoter sequence of the rice Os03g01700 gene, that is, the nucleotide sequence between upstream -1--3500bp from the ATG of the gene, and the promoter sequence can be transgenic in rice The regulatory system drives the specific expression of target genes in roots. The invention also relates to the application of the sequence in genetic engineering and genetic improvement. Background technique [0002] The successful application of plant genetic engineering technology not only requires a large number of promoters to be regulated at different levels, but also requires promoters suitable for different plant backgrounds, different organs, tissues, and transgenic types to avoid adverse effects during the transgenic process. For a long time, the non-plant endogenous constitutive promoter, which comes from cauliflower mosaic virus (cauliflowe...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113
Inventor 吴平李援亚刘绍军张帆刘于张为民
Owner ZHEJIANG UNIV