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Application of thinopyron elongatum in formulating low-celiac syndrome antigen encoding DNA wheat

A technology of Thinium elongatum and celiac disease, applied in the field of application of Thinium elongatum in the creation of low celiac antigen coding DNA wheat

Inactive Publication Date: 2011-06-01
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Aiming at the problem that existing wheat contains a large amount of celiac disease antigens, the purpose of the present invention is to provide a new application of E. elongatum, that is, the application of E. elongatum in creating low celiac disease antigen-encoding DNA wheat

Method used

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  • Application of thinopyron elongatum in formulating low-celiac syndrome antigen encoding DNA wheat
  • Application of thinopyron elongatum in formulating low-celiac syndrome antigen encoding DNA wheat

Examples

Experimental program
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Embodiment 1

[0027] Example 1 Using the Homologous Sequence Method to Screen the Wheat Distant Grass with Low Celiac Antigen Coding DNA

[0028] Take the young leaves of Echinopsis elongatum, and use the CTAB method to extract DNA. The primers are P1: 5'-ATG AAG ACC TTTCTC ATC CT-3', P2: 5'-TCA GTT AGT ACC GAA GAT GCC-3', PCR The enzyme used in the reaction is LA Taq (TaKaRa); cycle conditions: initial denaturation at 94°C / 3min, followed by 30 cycles (denaturation at 94°C / 40s, annealing at 55°C / 1min, extension at 72°C / 2min), and final extension at 72°C / 10min, the PCR results are shown in Figure 1.

[0029] PCR products were electrophoresed on 2.0% agarose gel and recovered (TIANGEN DNA recovery kit). The PCR recovered product was connected to the pMD18-T vector, and then transformed into E.coli DH10B competent cells. The method of cloning and transformation refers to the molecular cloning experiment guideline (Sambrook et al, 1989). The identified positive clones are sequenced bidirecti...

Embodiment 2

[0033] Example 2 Obtaining Wheat Somatic Hybrids Using Asymmetric Somatic Hybridization

[0034] 1) Acquisition of amphipathic embryogenic cells: the recipient material is wheat, and the donor material is distantly related grasses containing low celiac antigens, such as diploid and decaploid Henoptera elongatum. The immature embryos of the recipient and donor are in MS The callus was induced on the medium, and after subculture, the embryogenic callus was selected as the recipient.

[0035] 2) Collection of recipient and donor protoplasts: Take the materials that have been subcultured for 2-5 days, chop them up, use enzymatic hydrolysis technology, that is, at 25°C, free for 3-5 hours, filter with 300-500 mesh stainless steel mesh, Centrifuge at 500rpm for 5min to collect protoplasts.

[0036] 3) UV treatment of donor protoplasts: after washing the collected donor protoplasts with washing solution, adjust the density to 1×10 5-6 / mL, spread on the lower part of a petri dish w...

Embodiment 3

[0049] Example 3 Screening of low-celiac antigen-encoding DNA wheat somatic hybrids

[0050] The method of implementation is the same as the steps and methods in Example 1.

[0051] DNAs encoding four celiac antigens in hybrid II-12 (EU018336-EU018362) and parental wheat (Jinan 177) (EU018268-EU018299) and decaploid Echinopsis elongatum α-gliadin gene (EU018300-EU018335) See Table 1 for comparison.

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Abstract

The invention discloses an application that low celiac disease epitope coding DNA wheat can be created using couch grass with long ear. The application screens out distant grass of couch grass with long ear for low celiac disease epitope coding DNA wheat using a homologous sequencing; gene group or DNA of distant grass part is transferred into wheat using somatic cell hybridization technology before low celiac disease epitope coding DNA wheat somatic hybrid is screened out using the homologous sequencing. The somatic hybrid can be used for breeding work of low celiac disease epitope wheat and lay the foundation for food problem during therapy period for celiac disease patients.

Description

technical field [0001] The present invention relates to the application of Elongatum elongatum, in particular to the application of Etnoxypyronium elongatum in creating DNA wheat with low celiac disease antigen coding. Background technique [0002] Celiac disease (CD for short) is a systemic disease mainly celiac diarrhea mediated by the immune system after ingestion of gluten by genetically susceptible individuals. The incidence of this disease is higher in Europe and America, and the lowest in Africa. Gluten food is the pathogenic factor of celiac disease. In addition, individual genetic susceptibility, immune response and environmental factors affect the occurrence of the disease and complications. Gliadins (α-, β-, γ-, ω-) in gluten are the main pathogenic plant protein antigens, especially α-gliadins are toxic to the small intestinal mucosa. Studies have shown that: α-gliadin contains four celiac antigens, namely glia-α (QGSFQPSQQ), glia-α2 (PQPQLPYPQ), glia-α9 (PFPQP...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29A01H1/02C12N15/05
Inventor 陈凡国夏光敏张尚立陈梦竹赵峰
Owner SHANDONG UNIV
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