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Plant root hair development related protein TaRHD6, and coding gene and application thereof

A technology of encoding genes and genes, applied in the direction of plant gene improvement, application, plant peptides, etc., to achieve the effect of increasing root hair length, increasing plant biomass, and increasing root hair length

Active Publication Date: 2013-10-30
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Wheat, as the third largest food crop in my country, plays an important role in agricultural production, and the genes involved in the development of root hairs have not been reported yet

Method used

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  • Plant root hair development related protein TaRHD6, and coding gene and application thereof
  • Plant root hair development related protein TaRHD6, and coding gene and application thereof
  • Plant root hair development related protein TaRHD6, and coding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Embodiment 1, the cloning of wheat TaRHD6 gene

[0045] The root system of Yangmai 158 that had been hydrocultured for 8 days was extracted by Trizol method to extract total RNA, and after purification, it was reverse-transcribed with M-MLV reverse transcriptase to obtain cDNA. Using this cDNA as a template, PCR amplification was performed with primers L: 5'-TCACAGAGCCTAGCTAGCTAC-3' and R: 5'-TCTGATCTTACACGTGCCGC-3'.

[0046] PCR reaction system (30μl): dH 2 O 5.4 μl, 2×GC buffer 15 μl, 10 mM dNTPs (2.5 mM each) 0.6 μl, 100 U rTaq (Takara) 0.3 μl, each primer 3 μl, DNA template 3 μl.

[0047] PCR reaction conditions: 94°C, 5min; 35 cycles: 94°C, 30s, 59°C, 30s, 72°C, 1min; last extension at 72°C for 10min.

[0048] The obtained PCR amplification product was electrophoresed in 1% agarose gel, and a fragment of about 1 kb was recovered, connected to pEGM-T vector (Promega) and transformed into Escherichia coli DH5α strain for sequencing. Sequencing results show that a ...

Embodiment 2

[0049] Embodiment 2, the construction of recombinant plant expression vector

[0050] Using the site-specific recombination of Gateway technology, the target sequence is transferred to the expression vector modified by Gateway (ie, the target vector) through the two-step reaction of BP and LR. Gateway technology is based on the well-researched λ phage site-specific recombination system (attB×attP→attL×attR). The two reactions of BP and LR constitute the Gateway technology ( figure 1 ). The BP reaction utilizes a recombination reaction between an attB DNA fragment or expression clone and an attP donor vector to create an entry clone. The LR reaction is a recombination reaction between an attL entry clone and an attR destination vector. LR reactions are used to transfer a sequence of interest to one or more destination vectors in parallel reactions. The donor vector used in this example is pDONR TM 221, the expression vector is pB2GW7.0, and its structure diagram is as foll...

Embodiment 3

[0076] Embodiment 3, wheat TaRHD6 gene transformation Arabidopsis

[0077] 1. Construction of recombinant Agrobacterium

[0078] Take the recombinant plant expression vector pB2GW7.0-TaRHD6 prepared in Example 2 to transform Agrobacterium tumefaciens GV3101 competent cells, and select and culture in YEB solid culture containing 50 μg / ml spectinomycin and 100 μg / ml rifampicin at 28°C Pick the positive clone and carry out the plasmid, and carry out PCR detection with the primer L and R of the full-length cDNA sequence of the amplified wheat TaRHD6 gene in embodiment 1; The recombinant Agrobacterium tumefaciens was named GV3101 / pB2GW7.0-TaRHD6.

[0079] 2. Obtaining transgenic Arabidopsis

[0080] 1) Vernalize the wild-type seeds of Arabidopsis thaliana Col-0 (Arabidopsis thaliana Columbia) at 4°C for 72 hours, sow them in MS medium at 22°C / 18°C, 15h light / 9h dark, and humidity Cultivate in 60%-70% of the culture room, and transplant it into a planting pot mixed with nutrient so...

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Abstract

The invention discloses a plant root hair development related protein TaRHD6, and a coding gene and application thereof. The protein disclosed by the invention is derived from wheat and named TaRHD6, and is composed of amino acid sequence disclosed as Sequence 2 in the sequence table. The experiment proves that compared with a wild Arabidopsis thaliana plant under the same conditions, the phenotype of a T3-generation homozygous transgenic plant obtained by transforming Arabidopsis thaliana with a recombinant expression vector pB2GW7.0-TaRHD6 of a DNA (deoxyribonucleic acid) molecule (the coding gene of the protein) disclosed as Sequence 1 in the sequence table has obviously increased root hair length; and under normal and nutrient stress conditions, the plant types and fresh weight of the overground plant are obviously increased. The invention has important meanings in the aspects of increasing the plant root hair length and further enhancing the biological yield of the plant.

Description

technical field [0001] The present invention relates to a plant root hair development-related protein and its coding gene and application in the field of biotechnology, in particular to a plant root hair development-related protein TaRHD6 and its coding gene and application. The protein TaRHD6 is derived from wheat and has the function of regulating plant root hair length, thereby regulating the function of plant biomass. Background technique [0002] Root hairs are single-celled, tubular protrusions formed by the outgrowth of specific epidermal cells near the root tip. The existence of root hairs significantly increases the surface area of ​​the root, which helps to improve the stability of the root in the soil, the interaction between the root and microorganisms, and the relationship between the root and the root. Absorption of soil nutrients, it is the most active organization for roots to absorb water and nutrients. Studies have shown that plants with longer root hairs ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21C12N7/01A01H5/00
Inventor 倪中福孙其信韩瑶姚颖垠彭惠茹
Owner CHINA AGRI UNIV
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