Gene for controlling arabidopsis trichome development and coded protein and application thereof

A kind of epidermal hair, Arabidopsis technology, applied in the field of plant genetic engineering

Inactive Publication Date: 2012-04-11
SHANXI ACAD OF AGRI SCI GARDENING RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the development of epidermal hairs involves the joint action of many genes, and the regulatory modes of many genes are still unclear. Therefore, the study of the mechanism of genes related to epidermal hairs by linking phenotypic variation and gene function is ve...

Method used

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  • Gene for controlling arabidopsis trichome development and coded protein and application thereof
  • Gene for controlling arabidopsis trichome development and coded protein and application thereof
  • Gene for controlling arabidopsis trichome development and coded protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Construction and screening of Arabidopsis thaliana activation tag mutant library

[0031] 1) Arabidopsis seeds were cold-treated at 4°C for 3 days to germinate neatly. Sow in humus soil containing 1 / 3 vermiculite for cultivation. The culture room is kept at a temperature of 22-24°C, a relative humidity of 80%, the light source is a fluorescent lamp, and the light intensity is 85 μmol.s -1 .m -2 , the light time is 16h;

[0032] 2) About 4-5 weeks, Arabidopsis begins to flower, which can be used for transformation;

[0033] 3) Pick a single colony of Agrobacterium containing the pDSK15-11 plasmid in YEP liquid medium (composed of yeast extract 10g / L, peptone 10g / L, sodium chloride 5g / L, pH7.0Gen 100mg / L, Carb 50mg / L, Rif 50mg / L), 28 ℃, shaking culture under 200rpm overnight; Transition to fresh YEP liquid medium containing antibiotics in a ratio of 1:50 and expand to the logarithmic growth phase (bacterial liquid OD600 is about 1.5); centrifuge at 5000×g...

Embodiment 2

[0038] Example 2: TAIL-PCR amplification of T-DNA flanking sequences and identification of T-DNA insertion sites

[0039] In the present invention, the amplification of the T-DNA flanking sequence adopts the method of TAIL-PCR (thermally asymmetric interleaved polymerase chain reaction), the PCR product is directly sequenced, and the T-DNA is found after comparing with the whole genome sequence of Arabidopsis thaliana Inserted between At4g01050 and At4g01060 of chromosome 4 of Arabidopsis thaliana, about 2000 bp away from the start codon of At4g01060, and its right border points to At4g01060. In order to further determine whether the mutant phenotype is caused by this T-DNA insertion, the present invention carried out genetic analysis on the offspring of the T2 generation of B11. Among the 31 plants of the T2 generation, 24 of them had the phenotype of no epidermis. , the 7 strains are normal, and the segregation ratio of phenotype and no phenotype is about 3.4:1, so it can al...

Embodiment 3

[0071] Example 3: RT-PCR detection of T-DNA insertion site flanking genes and identification of genes controlling Arabidopsis trichome development

[0072] In the present invention, because the T1 generation of B11 shows the phenotype of lack of epidermis, this indicates that the mutant phenotype is a dominant mutation, which is caused by the activation of the flanking gene by the activation tag. The present invention selects 4 genes in the range of 10kb upstream and downstream on both sides of the T-DNA insertion site, including upstream At4g01040 and At4g01050 and downstream At4g01060 and At4g01070, and the expression levels of these 4 genes are carried out by RT-PCR Semi-quantitative detection, such as Figure 6 As shown, the RT-PCR results showed that among the four genes, only At4g01060 was expressed in the B11 mutant, and the expression of the other three genes was not different between bzr1-1D and the mutant B11. This result indicated that the phenotype of the mutant w...

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Abstract

The invention relates to the field of plant gene engineering, in particular to a gene for controlling arabidopsis trichome development and a coded protein and application thereof. The nucleotide sequence of the gene has the sequences as shown in the SEQ ID NO:1, and the amino acid sequence has the sequences as shown in the SEQ ID NO:2. The gene plays an important role in regulating leaf and stem trichome development and is a negative regulator in trichome development. A mutant B11 with leaf and stem trichomes deleted is screened by a method of activating indexes. The T-DNA is found to be inserted between the No.4 chromosomes At4g01050 and At4g01060 of arabidopsis after the T-DNA flanking sequence is subjected to amplification, sequencing and database comparison by a TAIL-PCR (thermal asymmetric interlaced-polymerase chain reaction) method. The phenotype of the mutant is caused by overexpression of At4g01060, and the overexpression can cause reduction of leaf and stem trichomes. The study further perfects the regulation mode of trichome development, lays the theoretical and production practice foundations for cultivating new species of genetically modified crops and has important practical significance for agricultural production and crop improvement.

Description

technical field [0001] The invention relates to the field of plant genetic engineering, in particular to a gene for controlling the development of the epidermis of Arabidopsis thaliana, its encoded protein and its application. Background technique [0002] The development of epidermal hairs is a very useful model for studying the regulation of plant cell differentiation. Its morphogenesis is obviously different from that of the surrounding epidermal cells, and its origin and development are also easy to observe because of its specificity. Its main function is to increase the thickness of the epidermis to reduce heat and moisture loss and UV radiation, and to protect plants from insects and pathogens and some mechanical damage (Szymanski et al., 2000). [0003] In recent years, great progress has been made in the research on the molecular genetic control mechanism of Arabidopsis trichome development. Several genes are currently known to be involved in regulation, including G...

Claims

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Application Information

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IPC IPC(8): C12N15/29C07K14/415C12N15/84A01H5/00
Inventor 曹冬梅范喜英王云山康黎芳张超段九菊
Owner SHANXI ACAD OF AGRI SCI GARDENING RES INST
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