Rice seed glutelin glub-5 gene terminator and its application

A gene and transgenic plant technology, applied in application, angiosperm/flowering plants, introduction of foreign genetic material using vectors, etc., can solve the problems of insignificant improvement of related traits, time-consuming and labor-intensive, etc., to increase the added value of science and technology, and achieve large-scale applications. Prospects, effects of increasing expression and accumulation levels

Active Publication Date: 2011-11-30
INST OF BOTANY CHINESE ACAD OF SCI
View PDF0 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, less research has been done on other terminators that can enhance gene expression
Since some important agronomic traits and plant secondary metabolites are controlled by multiple genes, increasing the expression of a single gene has no obvious effect on the improvement of related traits
Time-consuming and labor-intensive transformation of multiple genes through traditional transformation methods such as repeated transformation or hybridization

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Rice seed glutelin glub-5 gene terminator and its application
  • Rice seed glutelin glub-5 gene terminator and its application
  • Rice seed glutelin glub-5 gene terminator and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1, the acquisition of rice seed glutelin GluB-5 gene terminator (tGluB-5)

[0039] According to the cDNA sequence of rice glutelin GluB-5 gene (GenBank number is AK107238), search the genome DNA sequence of glutelin GluB-5 gene from GenBank, the 497bp sequence after the stop codon is the rice seed glutelin GluB-5 of the present invention. 5 gene terminator (tGluB-5), design primers to amplify tGluB-5. To facilitate vector construction, restriction sites (underlined) were added to the primers respectively. The forward primer of tGluB-5 is tGluB-5SacF: 5′-AA GAGCTC ACCCAAGGCATTATACTAA-3'(Sac I), the reverse primer is tGluB-5EcoR:5'-A GAATTC AAACTTGGTGCCACGACACTG-3' (EcoR I).

[0040] A small amount of genomic DNA was extracted from the leaves of wild-type rice Kitaake (Qu et al., J. Exp. Bot. 2008, 59: 2417-2424) by CTAB method, using it as a template and tGluB-5SacF and tGluB-5EcoR as primers, PCR amplified tGluB-5 sequence. The PCR reaction program was...

Embodiment 2

[0041] Example 2, Vector Construction and Transformation of Rice Seed Glutenin GluB-5 Gene Terminator (tGluB-5)

[0042] 1. Construction of tGluB-5 fusion GUS gene plant expression vector

[0043] Digest the pMD-tGluB-5 plasmid with Sac I and EcoR I, recover the 497bp restriction fragment of tGluB-5, and insert the fragment into pGluB-3-nos containing GluB-3 promoter and pGluC of GluC promoter respectively The stable transformation vector pGluB-3-tGluB-5 was constructed between the Sac I and EcoR I double restriction recognition sites of -nos (the schematic diagram of which is shown in image 3 Shown in A) and pGluC-tGluB-5 (the schematic diagram of its structure is shown in image 3 shown in B).

[0044] pGluB-3-nos and pGluC-nos were constructed according to the method described in the literature Qu et al., J.Exp.Bot.2008, 59: 2417-2424: the genomic DNA of rice Taichung 65 was used as a template, and

[0045] GluB-3 Forward Primer 5'-CCC AAGCTT ATTTTACTTGTACTGTTTAACC-3'...

Embodiment 3

[0058] Example 3, Functional Verification of Rice Seed Glutenin GluB-5 Gene Terminator (tGluB-5)

[0059] 1. T 0 Histochemical detection of GUS in transgenic rice

[0060] T of transgenic rice Kitaake / pGluB-3-nos and Kitaake / pGluB-3-tGluB-5 positive for PCR detection 0 The generation plants were subjected to histochemical staining, and the non-transgenic wild-type rice Kitaake plants were used as the control. The specific steps are: the T of transgenic rice Kitaake / pGluB-3-nos and Kitaake / pGluB-3-tGluB-5 0 Part of the leaves, roots, and stem tissues of the generation plant and the non-transgenic wild-type rice Kitaake plant were cut into small pieces; the seeds at the filling stage 17 days after flowering were cut longitudinally from the middle with a scalpel. Soak the treated sample in GUS staining reaction solution (0.1M sodium phosphate buffer (pH 7.0), 10mM Na 2 - EDTA (pH7.0), 5mM potassium ferricyanide, 5mM potassium ferrocyanide, 1.0mM X-Gluc, 0.1% Triton X-100), re...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses a rice seed glutelin GluB-5 gene terminator and application thereof. The terminator is a DNA (deoxyribonucleic acid) molecule from the following: 1) a DNA molecule composed of nucleotide sequence disclosed as Sequence 1 in the sequence table; 2) a molecule which is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% homological with the DNA defined in the 1); and 3) a molecule crossbred with the DNA sequence defined in the 1) or 2) under strict conditions. By cooperating with different promoters, the terminator disclosed by the invention can enhance the expression and accumulation level of exogenous gene in the target plant, lays foundation for researching seed quality improvement, molecular medicine farms and the like, and has wide application prospects.

Description

technical field [0001] The invention relates to a rice seed glutelin GluB-5 gene terminator and application thereof. Background technique [0002] The latest development of plant biotechnology not only realizes the improvement of traditional agronomic traits (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 . Most gramineous crops have the characteristics of high yield, low production cost, storage resistance, easy control of production scale, direct consumption, etc., and they have the ability of post-translational modification in vivo, so they become ideal carriers for the second generation of transgenic products. In recent years, the use of rice seeds to produce exogenous proteins and edible vaccines with health effects has developed rapidly, and has achieved great success. Such as vitamin A, glycinin which can l...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21C12N7/01A01H5/00
Inventor 曲乐庆李文静
Owner INST OF BOTANY CHINESE ACAD OF SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap